Winter ’15 Vol. 28 No. 1 bulletin - Science Forward · PDF filevol. 28 / no. 1 bulletin...

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bulletinWinter ’15 Vol. 28 No. 1

in this issue Allergy, Asthma, and the Gut

Germline Stem CellsScience Posses Fan Out

Super Resolution

Nobel-winning work provides a stellar

view of cellular detail

Cradle of LifeMystery and awe surround the genesis of life, whether observed

from afar or through the lens of a microscope. Even in the ovary of a fruit fly, as seen here, the beauty and wonder are undeniable.

The orb-like organ is where primordial germ cells, or PGCs (green), rub shoulders with special “intermingled cells” (red), contact that is

required for the germ cells to proliferate. Later in development, some of the PGCs will become germline stem cells—destined to start the process all over again—while the others will differentiate into eggs.

Learn more about how germline stem cells hold the key to survival of a species, including humans, in “Perfect Balance,” page 18.

4000 Jones Bridge Road Chevy Chase, Maryland 20815-6789www.hhmi.org

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12Immune cells will do anything within their power to do their jobs. Some, like this neutrophil, are driven to crawl, swim, or ooze through our bodies’ tissues and fluids to be a first responder at an acute infection or inflammation. Until recently, we couldn’t have known what these cells look like in action, as they morph from a sphere into a shape-shifting tangle of appendages and ruffled edges. But now, remarkable advances in imaging live cells offer a whole new view of biology.

HHMI Bulletin / Winter 2015

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The Essence of our BeingWe humans may think of ourselves as solid objects, all flesh and bone. But take a close look, and it’s clear our bodies are composed largely of oxygen and hydrogen. We are essentially ephemeral—akin as much to wind, water, and fire as to earth. No matter how heavy you feel, says biologist and author Curt Stager, you are more like a “porous froth of atomic Styrofoam” than a dense mass. Understanding the world in atomic terms can give us a richer appreciation, he says, for how we interact with each other and the environment around us.

What do atoms have to do with you? Everything. They were present and intimately involved when you and everyone you have ever loved—or hated—did everything that you and they have ever done. Every scent you’ve ever savored, every sight you’ve ever seen, every song you’ve ever enjoyed, every cry or sigh that ever passed your lips sprang from atoms at work within the atmosphere and the darkest recesses of your body. When you eat, the bodies of other living things become part of you. If you cut yourself, the wreckage of dying stars runs out in a stream of ancient atoms that triggered some of the most violent explosions in the cosmos. When you flush your wastes, you scatter the atomic echoes of lightning bolts and volcanoes into a global cycle that may some day return them to you, as unpleasant as that may sound. And whenever you grin, the sparkling of your teeth conceals the dim afterglow of nuclear fallout from Cold War bomb tests over the Pacific.

You are not only made of atoms; you are atoms, and this book, in essence, is an atomic

field guide to yourself. All you need in order to interpret your life in primal elemental terms is access to some of the latest scientific information, some new ways to reconsider your world in light of it, and an active imagination. In doing so you will begin to experience a revolution in self-awareness that is playing out on a larger scale around the world.

From Your Atomic Self by Curt Stager. Copyright ©

2014 by the author and reprinted by permission of

Thomas Dunne Books, an imprint of St. Martin’s

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ContentsWinter ’15 Vol. 28 No. 01

Web-Only Content

Follow a group of Michigan scientists as they face flaming logs and pits of mud in a Warrior Dash.

Get up close and personal with the porcupine-like projections of a HeLa cell.

See videos generated by the innovative microscopes at Janelia’s Advanced Imaging Center.

Go wild with a slideshow of Susan McConnell’s nature photography.

Travel to Russia to visit a summer school that gives talented Russian students a taste of life at the bench.

Departments president’s letter03 DiscoveryandPerseverance centrifuge04 WarriorforaDay05 SecondCalling benchreport06 MemorySwap08 ImmunityMeetsMetabolism10 TheProteinattheEndof theTunnel perspectives&opinions30 RethinkingGMORegulation32 Q&A–Howcanscientistshelp easesociety’sfearofGMOs? chronicle34 Science Education ImmersionintheLab36 Toolbox MicroscopesfortheMasses38 Lab Book Decisions,Decisions Cancer’sNicheProblem JammingJumpingGenes observations TheEssenceofourBeing

Features12 Defying Limits Fueledbyadesiretohelp biologists,EricBetzighas spentthelastdecadedevising waystobreakfreefromthe constraintsoflightmicroscopy— workrecognizedwiththe2014 NobelPrizeinChemistry.

18 Perfect Balance Germlinestemcellstreada finelinebetweenreplenishing themselvesandproducing spermandeggs.Theyjustmay bethemostimportantcells you’veneverheardof.

24 Power to the Posse Servingasbothalifelineand alaunchingpad,thePosse STEMprogrampromotes persistenceamongmulticultural collegestudents.

Defying Limits, page 12

www.hhmi.org/bulletin

Cover illustration:

Markos Kay

This paper is certified by SmartWood

for FSC standards, which promote

environmentally appropriate, socially

beneficial, and economically viable

management of the world’s forests.

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Contributors

Telephone (301) 215.8500 • Fax (301) 215.8863 www.hhmi.org© 2015 Howard Hughes Medical InstituteTheopinions,beliefs,andviewpointsexpressedbyauthorsintheHHMI Bulletindonotnecessarilyreflecttheopinions,beliefs,viewpoints,orofficialpoliciesoftheHowardHughesMedicalInstitute.

HHMI TRUSTEESKurt L. Schmoke, Esq., ChairmanPresident / University of BaltimoreJames A. Baker, III, Esq.Partner / Baker Botts. LLPAmbassador Charlene BarshefskySenior International Partner / WilmerHaleJoseph L. Goldstein, MDRegental Professor & Chairman Department of Molecular Genetics University of Texas Southwestern Medical CenterGarnett L. KeithChairman & CEO SeaBridge Investment Advisors, LLCFred R. LummisChairman & CEO Platform Partners, LLCSir Paul NursePresident / The Royal SocietyDame Alison F. Richard, PhDSenior Research Scientist & Professor Emerita Yale UniversityClayton S. Rose, PhDProfessor Harvard Business SchoolAnne M. TatlockRetired Chairman & CEO Fiduciary Trust Company International

HHMI SENIOR EXECUTIVE TEAMRobert Tjian, PhD / PresidentCheryl A. Moore / Executive V.P. & Chief Operating OfficerKathryn S. Brown / Head of CommunicationsSean B. Carroll, PhD / V.P. for Science EducationJennifer L. Farris / Senior Director, Campus ServicesHeidi E. Henning / V.P. & General CounselMohamoud Jibrell / V.P. for Information TechnologyNitin Kotak / V.P. & Chief Financial OfficerErin O’Shea, PhD / V.P. & Chief Scientific OfficerGerald M. Rubin, PhD / V.P. & Executive Director, Janelia Farm Research CampusKathy A. Wyszynski / V.P. for Human ResourcesLandis Zimmerman / V.P. & Chief Investment Officer

HHMI BULLETIN STAFFMary Beth Gardiner / EditorDana Cook Grossman / Story EditorJim Keeley / Science EditorNicole Kresge / Assistant Editor

ADDITIONAL CONTRIBUTORSElaine Alibrandi, Michelle Cissell, Heather McDonald Pentagram Design / DesignAllied Printing / Printing & Binding

Henry McCausland(“WarriorforaDay,”page4)livesinLondonandworksasaninternationalillustrator.Heenjoyspens,hishealth,sunshine,andbeingaskedtodrawpeoplerunning.

ACalifornia-basedphotographer,Leslie Williamson(“RethinkingGMORegulation,”page30)isbestknownforherportraitsofpeopleandtheirspaces,andforhertwobooks,Handcrafted Modern: At Home with Midcentury Designers and Modern Originals: At Home with Midcentury European Designers.Shetravelstheworldinsearchofcreativepeopleworkingonexcitingprojects.

Connecticut-basedwriterCathy Shufro(“ImmersionintheLab,”page34)alsotutorswritingatYaleCollege.A2012InternationalReportingProjectfellow,sheneverconsideredvisitingRussiauntilwritingherstoryforthisissue.Nowshe’sreadingthesatiricalnovelthatRussianscientistMikhailGelfandrecommended—Heart of a Dog,byMikhailBulgakov.

Markos Kay(coverand“DefyingLimits,”page12)isadigitalartist,illustrator,andlecturerbasedinLondon.Hisworkexploresthevisualizationandabstractionofthemicroscopicworldthroughcomputersimulations.Heisakeencollaborator,alwaysonthelookoutforscientistsinterestedinvisualizingtheirdata.

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President’s Letter

Discovery and Perseverancelastmonth,ericbetzig ,agroupleaderatHHMI’sJaneliaResearchCampus,traveledtoSwedentoaccepttheNobelPrizeinChemistry.Ericsharedtheawardwithtwootherscientists,StefanHell,oftheMaxPlanckInstitute,andWilliamMoerner,ofStanfordUniversity,fortheirremarkablecontributionstopushbeyondthelimitsoflightmicroscopy.ItwasawatershedmomentforEricandforallofusatHHMI.Afterall,JaneliawascreatedwithpeoplelikeEricinmind—scientistswhoareintellectuallydaringandcommittedtodiscovery,productivity,andperseverance.

EricjoinedHHMIinOctober2005,ayearbeforethedoorsopenedatJanelia.HesharedtheInstitute’svisionforthecampus:aplacewheresmall,interdisciplinarygroupsworkcollaborativelytobuildpowerfultoolsandpursuesomeofscience’smostprofoundquestions.Andhehelpedrecruitothertalentedscientists,includinghisclosefriendandPALMmicroscopecollaboratorHaraldHess.

AstheyandotherJaneliansbuilttheirlabsinthoseearlydays,theywerealsohelpingtoestablishtheprinciplesandculturethatnowcharacterizethisuniquecommunity.Fromtheoutset,sharingJanelia’sresourcesandtoolswiththebroaderscientificcommunitywaspartofourplan.Oneexampleofthatisourvisitingscientistprogram.WealsorecentlypartneredwiththeGordonandBettyMoore

FoundationtoestablishanAdvancedImagingCenter,whereselectscientistsfromaroundtheworldcanaccesspotentiallytransformativeimagingtoolsdevelopedatJanelia,includingthosedevelopedbyEric.

Myownworkhasbenefitedgreatlyfromsuper-resolutionimagingsystemslikePALMandEric’smorerecentlatticelightsheetmicroscope,whichcapturesthree-dimensionalimagesoflivingcellswithexceptionalclarityanddetail.Thesetoolshaveallowedmyteamtotrackthemovementandbindingactivityofindividualtranscriptionfactors,whichhelptranscribeDNAintoRNA,inlivinghumanoranimalcells.Wecandeterminehowlongaproteinneedstofindatargetsite,howmuchtimeitspendsatthatsite,andinwhatorderthemultipleproteinsthatmakeupthetranscriptionmachinecomplexareassembled.

Mylabhaslonghopedto“watch”transcriptionfactorsinactioninnormalanddiseasedcells.Wearestillearlyoninthisexciting

journey,butweknowthatsingle-molecule,single-cellimagingwillsignificantlyimpactstudiesoftranscriptionmechanisms,whichwillinturnhelpusunderstandhowgenesareexpressedduringanorganism’sdevelopmentandhowdiseasedisruptsthisprocess.

It’sworthnotingthatthe2006publicationonPALMwasthefirstjournalarticletocomeoutofJanelia.Today,asJaneliaapproachesits10thanniversary,thecampusincludessome800people,includingscientistsinfieldsrangingfromcomputationtoengineeringtoneuraltheory.Wehavemuchworktodo—andmorediscoveriesahead.

“From the outset, sharing Janelia’s resources and tools with the broader scientific community was part of our plan.”—roberttjian

Centrifuge4 Winter 2015 / HHMI Bulletin

Warrior for a Dayhotbutnot yetsweaty,thesmallgroupofscientistshadditchedtheirlabcoatsforamotleyassortmentofspandexandmicrofiberathleticgear(saveoneoutlierinanorangejumpsuit).Theystoodinavastfieldamongthousandsofsimilarlycladindividuals,stretchingandjogginginplace,exchangingnervousbanter,andeyeingtheothercompetitors.Overthenexthourorso,they’dtesttheirmettlenotbytryingtodesignthebestexperimentsandpublishthemostimpressivefindings,butbyleapingoverflaminglogs,wadingthroughpitsofmud,andscaling20-foot-tallcargonets.

TheharrowingfeatswerepartofaneventcalledtheWarriorDash.“Yourunlikecrazyandgoasfastasyoucan,”saysmolecularbiologistJamesBardwell,anHHMIinvestigatorattheUniversityofMichigan.BardwellandbiochemistUrsulaJakob,alsoamemberoftheMichiganfaculty,recruited11membersoftheirlabs—twopostdocs,fourgraduatestudents,andfiveundergrads—toentertheeventwiththem.Noneoftheirlabmateshadeverdoneanobstacleracebefore,butallweregame.“Itsoundedlikeareallyfun,healthy,team-buildingthingtodo,”recallsAdamKrieger,anundergradinJakob’slab.

“Ourlabshavedonethingstogetherbeforethis,”saysJakob.“ButIhavetosay,theWarriorDashwasdefinitelythebestexperiencebecausewewereallinittogether,juststrugglingtogetthrough.”

Billedasthe“world’slargestobstacleraceseries,”Warrior

Dashesareheldyear-roundandworldwide;theMichiganscientiststookpartinalateJulyeventinMountMorris,Michigan.AWarriorDashsendsparticipants—aka,warriors—onagrueling5Kcoursethrougha“battleground”strewnwithobstaclesbearingformidablenameslikeMuddyMayhemandDieselDome.Afterward,they’rerewardedwithamassivepartyrepletewithlivemusic,food,andplentyofbeer.

Theeventalsohasaseriousside.WarriorscanraisemoneyforSt.JudeChildren’sResearchHospital.AndmanyhundredsofmuddyrunningshoesdiscardedbyparticipantsaftertheracearegroundintoplaygroundsurfacingmaterialbyacompanycalledUSAgain.Since2009,warriorsat200-plusraceshaveraisedmorethan$10millionforSt.Judeanddonatedsome70,000pairsofshoes.They’vealsodevoured35,000poundsofturkeylegs.

BardwellandJakobbelieveeventsliketheWarriorDashhelpbuildacamaraderiethatcarriesovertotheirlabs.Attherace,the13membersof

theirgroupsplitintosmallersquadsthathelpedeachothernavigatethechallengingterrain.“Youreallyhavetoworktogether,”saysJakob.“Scienceisallaboutcommunicationandcollaboration,andnotmuchdifferentfromanobstaclecourse.”

Bythetimetheycrossedthefinishline,thewarriorswerebruised,tired,andgratefulforthemassiveoutdoorshowerserectedbyraceorganizers.“You’retotallycoveredinmud,”saysBardwell.“Everybody’stakingoffshoesbecausethey’reallmuddy.Andeveryoneisfeelingreallygood.

“Theexperiencebroughtthelabtogether—there’snoquestion,”headds.“Whenyougothroughsomeeventthat’stough,youpulltogetherasateam.”—Nicole Kresge

For a GoPro video of the race, visit www.hhmi.org/bulletin/

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Second Callingit happenednearly adecadeago,butSusanMcConnellremembersthemomentwithtelephotoclarity:shewasstandingonthedeckofashipintheSvalbardarchipelago,halfwaybetweenmainlandNorwayandtheNorthPole,watchingapolarbearleapfromoneicefloetoanother.

Shepressedhercameratohereye,tryingtoanticipateeachjump.Herbicepsached.Herfingersweresonumbshecouldnolongerfeeltheshutterbutton.“Iwascoldtothecore,”saysMcConnell,aneuroscientistatStanfordUniversity,“andIrealizedthatI’dneverbeenhappierinmylife.”

ThoughMcConnellhadtraveledtoSvalbardseekingjustsuchasight,shewasn’tanespeciallyseriousphotographer.Infact,shewasambivalentaboutphotography.Lookingthroughalens,worriedMcConnell,woulddisconnectherfromthemoment.

Instead,theoppositeprovedtrue.ThechallengeofcomposingashotdeepenedMcConnell’sexperience,makingherawareofeachsubtleshiftinthebear’sposture,eachchangeinlightandmood.ShereturnedfromSvalbardwiththefiresofanewpassionstoked.

Herpassionforwildlife,though,hadbeentherefromthestart.McConnellwasanSu

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“animal-obsessedkid,”shesays,thekindwhokepthamstersandrabbitsandparakeets,whospentdayswatchingacolonyofwoodratsbesideherfamily’sPennsylvaniahome.SheidolizedJaneGoodallanddreamedofsomedayfollowingthegreatprimatologist’sfootstepsintoanAfricanjungle.

Asanundergraduateinbiologyinthelate1970s,however,McConnellfoundbehavioralstudiesunsatisfying.Sure,shecoulddescribeamonkeyyawningorcourtingamate—butwhatwashappeninginitsbraintoproducethebehavior?

Sobegananeurobiologycareerdevotedtounderstandingthedynamicsofneuronsinthecerebralcortex.McConnell’sfocusshiftedfromwildanimalstolaboratoryanimals,whereitwouldremainfortwodecades.ThencamethetriptoSvalbard.

Wildlifephotographyturnedintoasecondvocation,andMcConnellquicklygraduatedfromcapturingprettypicturestotellingstorieswithherphotographs:aboutameerkat

familystaringdownamaraboustorkoraboutelephantsrescuingababystuckinanabandonedwatertrough,aswellasaboutthelargerthemesofconservation.

Onaplanetwheresomanycreaturesareimperiledbytherapacityofacertainbipedalprimate,McConnell’snewmissionistoproduce“powerfulimagesthatconnectuswithournaturalheritageandstimulateacommitmenttoconservation,”shesays.HerworkhasbeenpublishedinSmithsonian andNational Geographic magazinesanddisplayedingalleries.

Yetshedoesn’tconsidertakingpicturestobeanescapefromthelaboratory.Rather,shesays,conservationphotographyandneurobiologyaretwotrunksthathavegrownfromthesameroot.

Bothcanbetracedbacktotheexperiencesofthatanimal-lovinggirl,andMcConnell,whothisyearreceivedfundingasanHHMIprofessortohelplifesciencestudentscommunicateresearchthroughart,wantstopassthatlovealong.

AtStanford,sheteachesconservationphotographytoundergraduates,helpingthemlearntotellstorieswithapurpose.“JustasmuchasI’mexcitedbymyownpictures,”saysMcConnell,“I’mexcitedbythephotographsmystudentstake.”—Brandon Keim

To see more of McConnell’s photos, go to www.hhmi.org/bulletin/winter-2015.

Memory SwapResearchersgaininsightintohowemotion-tingedmemoriesarestoredinthebrain.imagineyourselfon thewarm,whitesandofatropicalbeach.Asyoulistentothelappingwavesandinhalethesalt-scentedair,youformamemorythatcansummonafeelingofcontentmentforyearstocome.Butifadestructivehurricaneroarsinandruinsyourvacation,rememberingthatsamebeachinthefuturemightevokeanxietyorfear.Canyourewireyourmemorypost-hurricane,ensuringthatwhitesandandsaltairwillcontinuetoevokepositivefeelings?

Perhaps.“Emotionisintimatelyassociatedwithmemoryofpastevents,butthe[positive

ornegative]valenceofmemoriesisactuallyquitemalleable,”saysHHMIInvestigatorSusumuTonegawa.Heexplainsthatthebrainstoresamemory’scontextualinformation—detailsaboutwhereandwhenaneventoccurred—separatelyfromitsemotionalcontent.Thebraincircuitconnectingthosetwocomponentscanberewired,givingthememoryofaneventanewemotionalassociation,hesays.

There’sevidenceofthatmalleabilityinthesuccessofpsychotherapiesthathelppeopleovercomeanxietyordepressionbysummoningpositivethoughtstoreplacenegativeones.Tonegawa,aneuroscientistattheMassachusettsInstituteofTechnologywhostudiesmemoryinmice,wanteddirectevidenceofhowthebrainmakessuchaswitch.Soheandhiscolleagues,researchspecialistRogerRedondoandgradstudentJoshuaKim,devisedastrategytoreplaceananimal’sfearofaspecificplacewithapositivememoryoftheverysamelocation.

Theysucceeded.Whattheydiscoveredhelpsexplaintheconnectionsbetweendifferentcomponentsofmemoryandhowalteringthoseconnectionsmightreassignamemory’semotionalvalue.“We’veprovidedsomescientificbasisforalreadyongoingpsychotherapyandfoundatargetinthebrainforfuturetherapies,”hesays.

Tonegawaandhiscolleaguesfirstcreatedfalsememoriesinmicein2013.Theyplacedmiceinasafeenvironment,thenmadethebraincellsstoringthememoryofthatplacesensitivetolight,afeaturetheresearcherscouldusetoactivatethecells.Whentheyswitchedonthosecellsinamouseexploringaperilousnewspace,itcausedthemousetofeartheoriginalenvironment.Sotheyknewtheycouldattachemotiontoamemorythatpreviouslyevokedneitherfearnorpleasure.Butonceananimalhaddevelopedfearofaplace,coulditsmemoryofthatplacebemadepleasurableinstead?

Tofindout,Tonegawa’steamplacedmalemiceinachamberwhosefloordeliveredamildshock.Thescientistshadpreviouslyintroducedalight-sensitiveproteinintotherodents’braincellsthatstoredinformationaboutthedangerousplace.Asintheir2013experiments,theydidthisbylinkingproductionoftheproteintoagenethatis

switchedonasmemoriesareencoded,thusrestrictinglightsensitivitytotheprecisecellsthatstoredthenewlyformedmemory.Theyfurtherrestrictedthelight-sensitiveproteintooneoftworegionsofthebrain—thehippocampus,wherecontextualinformationaboutamemoryisstored,ortheamygdala,whichhousesamemory’semotionalcontent.Thentheyremovedthemicefromtheuncomfortablechamber.

Afewdayslater,thescientistsartificiallyreactivatedthememorybyshiningalightontheanimals’brains.Themicestoppedtheirexplorations,freezinginplace—typicalfear-basedbehavior.

Totrytooverwritethatfear,thescientistsplacedthemiceinanewenvironment.Insteadofelectricalshocks,pleasureawaited:thenewcageheldfemalemice.Asthemicemingled,theresearchersreactivatedthemalerodents’memory-storingneurons.Theyswitchedonthecellsresponsiblefortheoriginalfearmemoryonesetatatime—insomemice,theyactivatedcellsinthecontext-storinghippocampus;inothers,theytriggeredmemorycellsintheemotion-storingamygdala.Thentheytestedwhatemotionstheanimalsnowassociatedwiththeoriginalchamber.

Theresultswerestriking.Whenthescientistsreactivatedtheamygdalacomponentoftheoriginalmemoryduringthepleasurableencounter,themalemiceretainedtheirfear.Butwhentheresearchersreactivatedthememory-storinghippocampalcellswhilethemiceinteractedwithfemales,thememoryoftheoriginalchamberacquiredanew

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emotionalassociation.Nowthemicesoughtoutenvironmentsthattriggeredthememory.Whentheyputtheanimalsbackintheunpleasantenvironment,themiceexploredwithouthesitation.“Theoriginalfearmemoryisverymuchlost,”Tonegawasays.

Theresearchersfoundsimilarresultswhentheyswitchedtheemotionofamemoryintheoppositedirection,replacingapleasurablememorywithfear.

Bylocatingregionsofactivityinmicroscopeimagesoftheanimals’brains,Tonegawa’steamconcludedthatasetofhippocampalneuronsstorescontextualinformationaboutamemoryandconnectstoeitheroftwosetsofamygdalaneurons:oneresponsibleforpositivememory,theotherfornegativememory.Thememory-altering

experimentsrewiredthoseconnectionsaftertheyhadbeenestablished.

Thestudy,publishedSeptember18,2014,in Nature,confirmsthatbraincircuitsthatattachnegativeemotiontoamemoryarenotpermanent.Further,itidentifiesaspecificpartofthebrainasapotentialtargetfornewtreatmentsagainstdepressionandothermentalillnesses.

“Totargetdrugsoranyothertherapytoaparticularareaofthehumanbrainisachallenge,”Tonegawaacknowledges.“Butthetechnologydevelopssofast.It’squitepossiblethatthere’sawayofnoninvasivelymanipulatingcells’activityinhumans.Thesekindsofstudieswithanimalmodelsareprovidinginformationabouthowtotargetthoseprocedures.”—Jennifer Michalowski

“Emotion is intimately associated with memory of past events, but the [positive or negative] valence of memories is actually quite malleable.”—susumutonegawaJo

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Bench Report

Immunity Meets MetabolismLong associated with allergies and parasitic infections, a rare immune cell now appears to have a role in other bodily processes as well.w h e n r i c h a r d l o c k s l e y attendedgradeschoolinthe1950s,peanutbuttersandwicheswerealunchtimestaple.Yet“bythetimemykidswenttoschool,theywereoutlawed,”saystheHHMIinvestigatorattheUniversityofCalifornia,SanFrancisco.

Allergiestonutsandotherfoodssoared50percentbetween1997and2011.Thesymptoms—rashes,breathingtrouble,andevenlife-threateninganaphylaxis—areduetooverblowninflammatoryresponses.Asthma,anotherconditiontracedtohyperactiveimmunecells,isalsoontherise.

Thatposesaconundrum.“Whywouldweevolveanimmuneresponsetotortureourselves?”Locksleywondered.

Recentworkbythephysician-turned-scientistsuggeststheanswermaylieinunusuallymphoidcellsthatsensenutrientintakeinthegut.Accordingtoconventionalwisdom,immuneresponseslinkedtoallergicinflammationservetoprotectusfromintestinalworms.Afterall,thebranchof

theimmunesystemblamedforallergyandasthma—“type2immunity”—alsohelpsfendoffparasiticwormsknownashelminths.

ButLocksleysawholesinthislineofthinking.Forone,scoresofvertebrateanimalsinthewildhaveworms.“Theydon’tgetasthmaorallergy—butwedo,”hesays.

Plus,headds,“Indevelopingcountries,wherepeoplerunaroundbarefoot,100percentofthepopulationhasintestinalhelminths.Theycomefromthesoilandwater.”Infectedindividualshaveheightenedtype2immunityyetcan’tcleartheworms;diseasesymptomsoftendon’tshowupforyears,ifatall.

CluestotheconundrumcamewhenLocksleyconsideredadifferentvantagepoint:theworm’s.Coulditbethatwormstapintometabolicpathwaysthatallowthemtothriveinthehumangutfordecades?

ThejourneythatledLocksleytoexplorethispossibilitybeganinthe1980s.Asapostdoc,hebecamefascinatedwiththeparasite Leishmania.Inonemousestrain,Leishmaniainfectionswelledtheanimals’footpadsbutseemedotherwiseharmless.Yetinanotherstrain,theparasitemadethemicewasteawayanddie.Throughcelltransferexperiments,researchershadlearnedthatcertainlymphocytes—CD4-expressingTcells,or“Thelpercells”—determineifamouseresistsorsuccumbsto Leishmaniainfection.

“ThesameTcellscankillyouorhelpyou.Something’sdifferentaboutthem,andIwantedtofindoutwhat,”recallsLocksley.

ItturnsoutthatCD4Tcellsarenothomogeneous.TheseThelper(Th)cellsassumedifferentrolesgovernedbytheirreleaseofsignalingproteinsknownascytokines.Micethatproducemostly“Th2”cytokines—interleukin-4(IL-4)andinterleukin-13(IL-13)—succumbtoinfectionby Leishmania,Locksleyandcolleaguesdiscovered.Butmicethatmainlymake“Th1”cytokines,suchasinterferongamma,resistinfection.

Atthetime,scientiststhoughtthatsubsetsofCD4Tcellswerethesourceofthesecytokines.ButwhenLocksley’steamdeletedtheIL-4andIL-13genesinmouseCD4cells,theanimalscouldstillfightoffcertainworms.However,blockingthesecytokineswithantibodiesrenderedthemicedefenseless.Thissuggestedthatsomeothercelltypealsosuppliesthecytokines.

Tofindthemysterycells,theresearchersfluorescentlytaggedthesecytokinegenesinmicetoseewherethecytokineswereexpressed.Theydiscoveredthatararecell—type2innatelymphoid,orILC2—wasapotentsourceofIL-13andIL-5,arelatedTh2cytokine.

Unlikemostimmunecells,ILC2sdonotcirculateinblood.Instead,theyresideinskin,heart,lung,and,mostprominently,smallintestineandfattissue.“Thatchangedthegame,”Locksleysays.

Hislab’srecentdiscoveryofILC2’sroleinmetabolismchallengeslongstandingbeliefsaboutotherimmunecells,includingeosinophils.Implicatedasinitiatorsofallergicinflammationandinfightingworminfections,eosinophilscanalsoaccumulateinthesmallbowel—butnooneknewwhy.“Somesurvivalfactormustbedialedup,becauseotherwiseyouwouldn’tseethesecellsthere,”Locksleysays.“They’retoorare.”

Analyzingreportermice,histeamidentifiedthatsurvivalfactorasIL-5,acytokineproducedbyILC2sinthesmallbowel.TheresultsappearedinOctober2013in Nature.Earlierthatyear,LocksleyandcolleaguesreportedintheJournal of Experimental Medicine thatILC2ssustaineosinophilsinfattissue,too.

MorerecentworkfromLocksleyindicatesILC2salsohelpkeeplungshealthy.Histeamtriggeredasthmainmicebysprayingpurifiedchitinintotheirlungs.Chitin,thebiopolymerthatmakesinsectbodiesbrittle,isalsofoundinmoltinghelminthsandtheireggs,aswellasinothercommonurbanallergenssuchasfungiandmolds.Locksley’steamdeterminedthatchitinunleashesmolecularsignalsthatactivatelungILC2sinthemice.TurningontheseTh2pathwaysprovedessentialforrestoringlunghealth,theteamreportedinMarch2014in Immunity.

ThefindingscallintoquestiontheassumptionthatILC2s,andtheTh2immuneresponsestheyactivate,serveprimarilytowardoffparasiticworms.Instead,Locksleysays,itmightbethatwormsexploit“metabolicpathwaysthataregoodfortissuesinordertoexpeditetheirownsurvivalandreplicationovermanyyears.”

ByexploringthesignalsthatguideILC2sinmetabolismandotherbodilyprocesses,scientistsmaylearnhowtocalmasthmaandallergicreactions.PerhapsPB&JcanevenreturnasaschoollunchstandbyforLocksley’sgrandchildren.–Esther Landhuis

“The same T cells can kill you or help you. Something’s different about them, and I wanted to find out what.”—richardlocksley L

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Richard Locksley wants to get to the root of allergy and asthma.

Winter2015/HHMIBulletin

The Protein at the End of the Tunnel Scientists reveal the structure of a protein that performs the final step in an iconic and long-studied signaling pathway in cells.e m b r y o n i c d e v e l o p m e n t a n d cancermayseemwildlydifferent,butitturnsouttheyhavealotincommon.ARubeGoldberg-likecellsignalingpathwaythatscientistshavebeenstudyingfor30yearsholdsthekeytotheunlikelyconnection.

NewresearchfromHHMIInvestigatorLeemorJoshua-ToratColdSpringHarborLaboratoryhascharacterizedtheendpointinthatpathway.

OnesimilaritybetweencancercellsandtheundifferentiatedstemcellsofanembryoisamicroRNAmoleculecalledlet-7,whichisdestroyedinbothcases.MicroRNAspreventcertaingenesfrombeingexpressedandtranslatedintoproteins;thelet-7microRNAturnsoffgenesthatpromotecelldivision.Withoutlet-7,cellskeepdividing—allowingtheexpansionofembryoniccellsor,laterinlife,ofcancers.

AmicroRNAbeginsasastretchofself-complementaryRNAthatfoldsuponitselflikeVelcro,formingadouble-strandedmoleculewithaloopattheendcalledamicroRNAprecursor.Thisprecursoristhenprocessedtoforma22-nucleotidemicroRNAthattargetsprotein-codingmessengerRNAs,eitherpreventingtheirtranslationintoproteinsorleadingtotheirdegradation.Incancersandinstemcells,let-7doesn’tmakeittomaturitybecauseitsprecursorismarkedfordestructionbya“tail”ofuridinebases—theU’sinthegeneticalphabet.Anotherprotein,Dis3l2,recognizesthispolyUtailanddegradesit,alongwiththelet-7RNAprecursor.

Joshua-Tor’slabgroupusedx-raycrystallographytoidentifythestructureofDis3l2boundtoastringofU’s,likethosethattaglet-7fordegradation.AstheyreportedinNatureonOctober9,2014,herteamfoundthattheRNAfitsintoafunnel-shapedcavityinDis3l2.Withinthemouthofthefunnelandalongitsnarrowend,Dis3l2makesuridine-specificcontactswithlet-7’stail—notjustrecognizingthetailasRNAbut“reading”itsindividualletters.

“Seeinghowmanybasesarereadinarowisquiteunusual,”saysJoshua-Tor.OnceDis3l2makesitscontactsalongthestringofU’s,itbeginstobreakdowntheRNA,letterbyletter,untiltheentirelet-7precursorisdestroyed.“Onceitgetsgoing,itjustchewsitallup,”shesays.

A Path of Twists and TurnsThedemiseoflet-7istheendpointofacomplexsignalingpathwaysetinmotionbyaproteincalledLin28.

Thestoryofthatpathwaybeginsinthemid-1980s,intheMassachusettsInstituteofTechnologylabofH.RobertHorvitz,nowanHHMIinvestigator.Horvitz’spostdocVictorAmbroswaslookingforgenesthatalterdevelopmentintheroundwormCaenorhabditis elegans.InthemutantsthatAmbros(nowattheUniversityofMassachusettsMedicalSchool)andHorvitzwerestudying,wormseitherfellbehindorjumpedaheaddevelopmentally.Oneofthegenestheyidentifiedwaslin-28;wormsinwhichthegenewasdefectivejumpedaheadindevelopment.

Similarly,inbothmiceandhumans,theLin28proteinisabundantinembryoniccellsbutlessensthroughoutdevelopment,becomingundetectableinfullydifferentiatedcells.

In2000,let-7cameontothescenewhenGaryRuvkun,atHarvardMedicalSchool,reportedthatinworms,alossoflet-7hadaneffectoppositetoalossoflin-28:itcauseddifferentiatedcellstoreverttoearlierdevelopmentalstages.Ruvkunalsoidentifiedlet-7asamicroRNA—thesecondeverdiscovered(afterC. elegans’lin-4,whichAmbrosdiscovered).Let-7targetslin-28messengerRNA,repressingitsgeneactivity.Ruvkunpublishedasecondpaperin2000showingthatthelet-7microRNAisconservedacrossmostorganisms,includinghumans.

Lin28soonfoundapracticalapplication.In2007,scientistsreportedthatturningonfourgenes,including lin-28,inhumanfibroblastcellsreturnedthecellstoanembryonicstate—essentially,intostemcells.

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ButthoughLin28andlet-7wereknowntohaveoppositeeffectsondifferentiation,theirrelationshipwasunclear.Itwasin2008thatRichardGregoryandHHMIInvestigatorGeorgeDaley,atBostonChildren’sHospital,collaboratedtoshowatthemolecularlevelthatLin28represseslet-7maturation.

“Topreservethestemness,orpluripotency,ofthecell,”explainsJoshua-Tor,referringtostemcells’abilitytodifferentiateintomanycelltypes,“youhavetodownregulatelet-7,andthewaythecelldoesthatisbymakingLin28.”

Soonafterward,NarryKimatSeoulNationalUniversityreportedexactlyhowLin28turnsofflet-7:bypromotingtheadditionofthepolyUtail.Andby2009,theKimandGregorylabshadfoundtheenzyme,calledaTUTase,responsibleforaddingthetail.

A Rube Goldberg-like cell signaling pathway that scientists have been studying for 30 years holds the key to an unlikely connection between development and cancer.

AstheLin28/let-7pathwaywasbeingelucidated,evidencewasalsomountingforalinkbetweenthatpathwayandcancer.In2009,Daley’slabteamfoundLin28’stwohumanhomologsin15percentofhumancancers.Inaddition,overexpressionofLin28ledtotherepressionoflet-7and,consequently,theupregulationofthegrowth-promotinggenesthatlet-7normallykeepsincheck.

Thefindingshelptoexplaintheconnectionbetweencancerandembryonicdevelopment,saysDaley.“There’sayinandyangbetweentheLin28RNAbindingproteinanditsmaintarget,thelet-7microRNA,”heexplains.“Thisyin-yangiscarefullyorchestratedduringorganismalandtissuedevelopmentand[is]dysregulatedincancerformation.”

Thestemcell-cancerlinkwasstrengthenedfurtherwhen,in2013,GregoryidentifiedDis3l2astheproteinthatdegradesuridine-taggedlet-7inmousestemcells.ItturnsoutthatDis3l2’shumancounterpartismutatedinPerlmansyndrome,ageneticdiseasethatleadstofetalovergrowthandapredispositiontoacertainkidneytumor.ItisalsotheproteinwhosecrystalstructureJoshua-Tordetermined.

ResearchersmaynowwanttoseeifDis3l2degradesuridine-taggedRNAmoleculesotherthanlet-7.“It’sunlikelythatthisenzymeexistssolelytodegradetheprecursortolet-7,”Gregorysays.“TheremustbesomeotherrelevantRNAsubstrates.”AsDaleynotes,manymessengerRNAmoleculeshavepolyUtails.CouldDis3l2actonthem,too?“There’satleasttheprovocativepossibility,”hesays.—Ashley P. Taylor

Leemor Joshua-Tor’s lab used x-ray crystallography to identify the structure of Dis312 bound to a string of uridine bases.

Fueled by a desire to help biologists, Eric Betzig has spent the last decade

devising ways to break free from the constraints of light microscopy—

byjennifermichalowski

defying limitswork recognized

with the 2014 Nobel Prize in Chemistry.

illustrationbymarkoskay


Eericbetzigis aphysicistandanengineer:hethinksintermsoflightwavesandenergy,andwhenhetinkersinthelab,itiswithlasersandmirrorsandbeamsplitters.He’sthefirsttoadmitthatheisnobiologist.Butlately,hislabattheJaneliaResearchCampushasbeenbustlingwithexperimentalbiologistswhoarrivetotingvialsofcellsandleavewithgloriouslydetailedvideosofmolecularlifeinmotion.Thosescientistsreturntolabsallaroundtheworldwiththeirmindsracing,fullofideasabouthowBetzig’snewestinvention—amicroscopethatletsuserscollecthigh-resolution,three-dimensionalimagesoverprolongedperiods—mighttransformtheirresearch.

It’snotthefirsttimebiologistshaveclamoredtogettheirhandsononeofBetzig’sgroundbreakingimagingtools.NewtechnologieshavestreamedsteadilyoutofBetzig’slabashehasresolutelytickedoffitemsontheambitiousto-dolistthathelaidoutwhenhecametoJaneliain2005:fastermicroscopes,gentlermicroscopes,microscopesthatlookdeeper,andmicroscopesthatrevealmoredetail.Nineyearslater,hesays,“I’mgettingclosetodone.”

Biologistsareaccustomedtodesigningtheirexperimentsaroundthelimitationsofavailabletools.Forfluorescencemicroscopy—themethodofchoiceforfindingandfollowingspecificmoleculesinsidecells—thathasinthepastmeantchoosingbetweenimagingspeed,detail,orthewell-beingofthesample.ButBetzighasalwaysbeeneagertohearabouthiscolleagues’frustrations.“Ifyouknowtheproblem,that’shalfthebattletocomingupwiththesolution,”hesays.“That’sthesourceofcreativity.”Betzighasspentthepastdecadeapplying hiscreativitytofindingwaysaroundtheusualimagingtrade-offs,thusgivingbiologiststhefreedomtoapply theircreativitytoinvestigatingthequestionstheyhopetoanswer.

InOctober,Betzig’screativitywasrecognizedwiththe2014NobelPrizeinChemistry;hesharestheprizewithStefanHelloftheMaxPlanckInstituteforBiophysicalChemistryinGermanyandWilliamMoernerofStanfordUniversity.Theprizecommitteelaudedthethreescientists

fortheirdevelopmentofsuper-resolvedfluorescencemicroscopy—methodsofvisualizingobjectssosmallthat,untilrecently,distinguishingthemwithalightmicroscopewasconsideredafeatthatwoulddefythefundamentallawsofphysics.

Living ColorBetzighelpedlauncharevolutioninsuper-resolutionmicroscopyin2006,whenhedevelopedamethodthatheandhiscollaboratorHaraldHesscalledphotoactivatedlocalizationmicroscopy,orPALM.Thetechniquecreatesstunninglydetailedimagesofcellsbytakingadvantageoffluorescentlabelingmoleculesthatcanbeswitchedonandoffwithapulseoflight.InPALM,asamplelabeledwiththesefluorescenttagsisimagedmanytimes,withasmallsubsetofthefluorescenttagsswitchedoneachtime.Becausejustasmatteringofmoleculesisglowingintheimage,eachonecanbepinpointedwithprecision.Compilingthousandsofimagesyieldsapictureinwhichnearlyallfluorescentlylabeledmoleculesshowupasindividualanddistinct.

TheJaneliacampus—wherebothBetzigandHessarenowgroupleaders—wasstillunderconstructionwhenBetzigfirstlearnedofthefluorescentprobesthatwouldmakePALMpossible.SoheandHessbuiltaprototypeofthemicroscopeinHess’livingroominLaJolla,California.Later,theysetupshopinatraileronthecampusoftheNationalInstitutesofHealth,wheretheyworkedwithcellbiologistJenniferLippincott-Schwartzandotherstogeneratethemicroscope’sfirstspectacularlyrevealingimagesofcellsatwork.“Thefirsttimeweputadilutesuspensionofthese[fluorescent]moleculesonthesurface,andweturnedonthephotoactivatinglight,weknewwehadit,”heremembers.“Itfeltlikeflippingaswitch.”

BetzigandpostdoctoralresearcherHariShroff“livedandbreathedPALM”duringhisfirstfewyearsatJanelia,addingfeaturesthatmadethetechniquemorepowerful,suchastheabilitytodetectmultiplecolors—meaningmorethanonekindofproteincanbetrackedatonce—andthepossibilityofimaginglivingcells.ButBetzigknewthemethodhaditslimits.“It’stooslow,andyouthrowtoomuchlightatthecells,”hesays,explainingthatgeneratingasinglePALMimagesubjectssamplestotoxiclightexposuretimeandtimeagain.Becauseofthat,Betzigrealized,theprinciplesofPALMwouldnevertranslateintowhatbiologistsreallycraved:thesameextraordinaryresolutioninthree-dimensionalimagesoflivingcells.

Withaspateofotherlabsbythenworkingtoenhancesuper-resolutionmicroscopy,Betzigdecideditwastimetomoveon.“Everybodyandhiskidsisterwasdoingsuper-resolutionthen.Youhavetogototheareaswhereotherpeoplearen’t,”hesays.“Thosearethemostfertileareastofindsomethinguseful.”

Fertile FieldsHesawplentyofopportunities.NowthathewasatJanelia,minglingdailywithscientistsworkingattheforefrontofneuroscienceresearch,hebecameintentonimprovingtheircapabilitytolookinsidethebrain.Toinvestigatehownervecellsmediatethoughtsandactions,scientistsneed

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visualaccesstoallpartsofthebrain—butmostmicroscopescan’tseeveryfarbeneaththesurface.That’sbecausetheinsidesofcellsbendlightinunpredictableways.Thefartherlighttravelsthroughtissuetothemicroscope,themoredistortedanimagebecomes.

“Iknewthattheseguysprobablydidn’trealizehowcrummytheirimaginggetsastheystartpeeringintofliesandmice,”Betzigsays.Butheknewtherewaslotsofroomforimprovement.Healsoknewthatastronomershadalreadysolvedasimilarproblem.Turbulenceinourplanet’satmospheredisruptslightasittravelstotelescopesonEarth,muchthesamewaylightbendsandbouncesasitpassesthroughbiologicaltissue.Tocorrectthesedistortions,astronomersshinealaserintotheskytowardtheobjecttheywanttoobserveandmeasurehowtheatmospheredistortsthelightfromthis“guidestar”asitreturnstoEarth.Thentheyusethosemeasurementstocancelouttheatmosphericaberrationsintheimagestheyseethroughtheirtelescope.

BetzigandNaJi,apostdoctoralresearcherinhislabwhohassincebecomeaJaneliagroupleader,mimickedthisstrategybyinsertingafluorescentbeadintobraintissuetoactasabiologicalguidestar.Byimagingthatguidestar,theycancalculatethecorrectionsrequiredtobringintosharpfocustheblurrystructuresinimagesofthesurroundingtissue.

Atthesametime,Betzigwasthinkingaboutanotherproblemthatlimitedbiologists’abilitytoimagelivingcells:

“If you know the problem, that’s half the battle to coming up with the solution. That’s the source of creativity. ”—ericbetzig

Eric Betzig enjoys helping biologists answer challenging questions.


toomuchlight.Lightisessentialtoactivatethefluorescencethatmakeschemicallytaggedmoleculesvisibleunderanopticalmicroscope,butitisalsotoxictocells.What’smore,itburnsoutfluorescentmolecules,solabeledmolecules’signalsfadeovertime.Becauseofthat,Betzig’sbiologistcolleaguestoldhim,theycouldn’talwaysmonitorcellularprocessesforaslongastheywouldlike,andtheyoftenhadtosacrificedetailorspeedtoprolongthelivesoftheircells.

Betzigsayspartoftheproblemisthat,althoughamicroscope’sobjectivelensfocusesonjustoneplaneofasampleatatime,mostmicroscopesshineabeamoflightallthewaythroughtheirsamples,bombardingout-of-focusregionswithnoimagingpayoff.AtaJaneliaconference,BetziglearnedthatErnstStelzer,nowattheBuchmannInstituteforMolecularLifeSciencesinFrankfurt,Germany,waslimitinglightexposurebyimagingwithasheetoflightinsteadofabeam.“Itwasanelegantlysimplesolutiontotheproblemofphotodamageandout-of-focusbackground,”saysBetzig.

“Thebestideasarethesimpleones,”headds.Thiswasonehethoughthecouldadaptforhigh-resolutionimaging.Stelzer’stechniqueprolongedtheperiodhecouldmonitortheactivitiesofwholecells,butthelightsheetsheusedweretoothicktorevealcells’innerworkings.

Thethicknessofthelightsheetcouldnotbereducedwithoutsignificantlyreducingthemicroscope’sfieldofview,soBetzigcameupwithadifferentilluminationstrategy:alongbutnarrowbeamoflightcalledaBesselbeamthatcouldbesweptacrosstheimagingfieldtocreate

anextremelythinvirtualsheetoflightbettersuitedforimaginginsidesinglecells.

By2011,Betzigandtwomembersofhislab,LiangGaoandThomasPlanchon,haddevelopedamicroscopethatcollectsunusuallydetailedimageswithminimaldamagetolivingcells.Cellbiologistswereelatedandimmediatelybeganharnessingthetechniquetomonitordevelopmentalprocessesingrowingembryos,towitnesssubtleshapechangesascellsmovedinthreedimensions,andtofollowotherfast-movingorprolongedprocesses.Still,Betzig’steamsuspectedtheycoulddomoretoamplifytheirmicroscope’simpact.

DuringtheBesselbeammicroscope’sdevelopment,Betzighaddiscoveredthatcellsstayhealthierwhenthelighttheyaresubjectedtoisspreadout,reducingitspeakintensity.Hehadfirstnoticedtheimprovementwhenhisteamsplititsbeamintosevenpartstospeedupimaging.Thatwasasurprisebenefit,anditremindedBetzigthatheandJi,workingwithJaneliaGroupLeaderJeffMagee,hadhadasimilarsuccessafewyearsearlierwhentheyreducedthedamagecellsexperiencedduringadifferentimagingtechnique,aprocessknownastwo-photonmicroscopy,bybreakingupthemicroscope’spulsesoflightintosub-pulses.“Isaid,‘That’sexactlywhathappenedwiththetemporaldivisioninthepulsesplitter—soofcoursewehavetospreadtheenergyout.’”

Soatthispoint,Betzigwonderedwhethersplittingthebeamstillmoremightboostthebenefits.

EachBesselbeam’scentralcoreissurroundedbyconcentricringsofweakerlight,andhewasconcernedabouthowthosemightinteractwhenthebeamsbegantocrowdoneanother.ButwhenhemodeledtheinteractionsamongalargefieldofBesselbeams,hismodelpredictedcertainarrangementswheretheundesirablelobesoflightwoulddestroyoneanother.Ifhecouldcreatethosepatterns,energywouldbespreadout,thealreadylowlevelsoflightexposurewoulddropfurther,andthepatternwouldlenditselftoenhancingspatialresolutionwithatechniquecalledstructuredillumination.“Whatyougetissortofatriplewin,”Betzigsays.

Whole New WindowTenyearsearlier,beforehelandedthejobatJanelia,Betzighadbeeninneedofabigidea.AfterseveralearlysuccessesatAT&T’sBellLabs,wherehewasthefirsttoimagesinglefluorescentmoleculesatroomtemperatureinsteadofinextremecold,hehadbecomefrustratedwithacademicscience.HeleftBellLabsandspentthenextsevenyearsleadingresearchanddevelopmentathisfather’smachinetoolcompanyinMichigan.Butthatdidn’tholdhisinterestforever.Hewasreadytoreturntothelab—butheneededtoconvincesomeonetohirehim.“Ifigureditwasmylastchancetomakeascientificcareer,”hesays.

Hisinstinctwastodelveintosomethingtotallynew.Butwhenhelearnedthatbiologistscouldnowtagspecificmoleculesinsidelivingcellswithfluorescentproteins—apowerhe’dlongedforduringhisBellLabdays—heknewitwastimetorenewhisfocusonmicroscopy.HeretreatedtoacabininMichigantoimmersehimselfintheoretical

Two-photon imagining of living zebrafish with adaptive optical correction (right).

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physicsanddevisedaschemetolimitlightdamagetocellsduringimaging.Thetechniqueheproposedinvolvedilluminatingsampleswithamassivethree-dimensionalarrayoflightfoci.He’dneverbuiltthat“opticallattice,”ashecalledit,butnowitwastimetoreturntotheidea.

“Irealizedthatthe‘magicperiods’[ofthetheoreticalBessel-beamlightfield]weretheperiodsthatwerepredictedbymyoptical-latticetheory10yearsago,”Betzigsays.Soheusedthattheorytomapoutthelightpatternsheexpectedwouldbeoptimalforimaging.OncepostdoctoralresearchersKaiWangandBi-ChangChenfiguredouthowtobestgeneratethosepatterns,theirmicroscopebeganproducingthree-dimensionalimagesofcellswiththedetail,speed,andlowtoxicitythatthey’dhopedfor.Themicroscopeissogentle,Betzigsays,that“therearemanycellsyoucouldlookatforever,in3-D.Inaway,it’sprettymuchthefinalsolutionforthat.”

Overthenextyear,Betziginvited30teamsofbiologiststovisitJaneliatoworkwithChenandfellowpostdocWesleyLegantonthenewmicroscope.Overthousandsofhours,theyworkedoutthekinkstobetterunderstandthetechnology’spotential,generatingterabytesofbreathtakingmoviesforeachgroup.Thosemovieswereunlikeanythescientistshadeverseen.Thelong,thinmicrotubulesthatgiveacellitsformandstructuralsupportcouldbeseenrapidlygrowing,shrinking,andregrowingascellsreshapedthemselvesandpreparedtodivide.Individualgene-activatingproteinscouldbefollowedinthreedimensionsastheydiffusedthroughcellsandboundtoDNA.Development-regulatinggrowthfactorscouldbemonitoredforhoursassmallclumpsofcellsbegantotransformthemselvesintomorecomplexorganisms.

“We’veworkedwithanewgroupofbiologistseveryweek,andeachtimeit’sanewadventure,”Betzigsays.“I’llneverbeabiologist,butIgetakickoutoftheartofit—outofthecrazinessofthecell.Itfeelslikewehaveawholenewwindowonnaturethatwedidn’thavebefore.”

Nowthattheyhavedemonstratedamiscellanyofapplicationsforthelatticelightsheetprocess,Betzigandhisteamareeagertocontinuedevelopingthetechnology.TheirhighestpriorityisintegratingarapidadaptiveopticstechniquethatBetzigandWangdevelopedlastyearforuseintransparenttissues.“We’regoingtotrytoextendthelatticelightsheetsowecan[image]throughtheentiretyoftransparentorganismslikethezebrafishor[theroundworm]C. elegansandhaveallthebenefitsofthenoninvasiveness,”heexplains.

Betzigdoesn’twantthatworktoimpederesearchers’accesstothecurrentlatticelightsheetmicroscope,sohisteambuiltaduplicateinstrumentforJanelia’snewAdvancedImagingCenter(see“MicroscopesfortheMasses,”page36).Thecenter’sdirector,Teng-LeongChew,saysthatmonthsbeforethelatticelightsheetmicroscopewasofficially

announced,itwasalreadythemostindemandoftheimagingcenter’smanycutting-edgetechnologies.Betzigandhisteamalsohelpedbuildtwomoreoftheinstruments,forlabsatHarvardandtheUniversityofCalifornia,SanFrancisco.Hehopesthelatticelightsheetprocess,whichhasbeenlicensedtoZeiss,willeventuallybecomestandardtechnologyinmostlabs.Butuntilthattimecomes,interestedresearchershavetheoptionofbuildingtheirowninstrument,byfollowingdetailedplansfromBetzig’slab.

HisteamreportedonthisnewmicroscopeinScienceonOctober24,2014—justweeksaftertheRoyalSwedishAcademyofScienceshadnamedBetzigaNobellaureate.Infact,BetzigworriedabitthattherecognitionforhisworkonPALMwasmisguided.“Thelatticewillbemylastingimpact,”hesays.“Myguessisthiswillbethehigh-watermarkofmycareer.”

Thoughworkonthelatticeisnotyetdone,Betzigexpectsthefinaloptimizationstoproceedsmoothly.“I’mjustdottingi’sandcrossingt’s,”hesays.“IprettymuchknowthatmostofthestuffI’mdoingrightnowisgoingtowork.”

ThatkindofstabilitymakesBetzigrestless.“It’stimetothrowthediceagain,”hesays.“It’stimetotrysomethingrisky—thekindofstuff[Janelia]isdesignedtosupport.”

To see videos from Betzig’s lattice light sheet microscope, go to

www.hhmi.org/bulletin/winter-2015.

“I’ll never be a biologist, but I get a kick out of the art of it—out of the craziness of the cell.”—ericbetzig

bymeganscudellari

Germline stem cells tread a fine line between replenishing themselves and producing sperm and eggs. They just may be the most important cells you’ve never heard of.

PerfectBalance

illustrationbyadamsimpson


Sstemcellsare amongthefewcellsinthebodywiththeabilitytogeneratenewcelltypes,butbecauseofthisdistinction,theyarecaughtinaprecarioussituation.Stemcellswalkatightropebetweenthetwomostpowerfulcellularforcesinthebody:theyareabletobothself-renew,producingmorestemcells,anddifferentiate,generatingspecializedcellswithspecificfunctions,suchasbloodorbonecells.Ifthebalancetipsinfavorofself-renewal,stemcellscanover-accumulate,insomecasesleadingto

cancer.Ifthebalancetipstowarddifferentiation,thepoolofstemcellscandisappear,leadingtothedeteriorationoforgans.

Nowhereisthattightropewalkmorevitalormoreperilousthaningermlinestemcells—thestemcellsthatultimatelyproducespermandeggs,orgametes.Whileallstemcellsself-renewandproducespecializedcells,germlinestemcells(GSCs)aretheonlyonesthatmakegametes,

whichareresponsiblefortransmittinggeneticinformationfromonegenerationtothenext.“They’retheonlycellsthatyoudon’tactuallyneedforyourownsurvival,butyoudoneedthemforthesurvivalofyourspecies,”saysRuthLehmann,anHHMIinvestigatorattheSkirballInstituteofBiomolecularMedicineatNewYorkUniversity.

Inthisway,GSCsaretheveryfoundationofaspeciesandmustwalkthattightropecarefully.“Inagermlinestemcell,justbeingpreciseisn’tenough,”saysYukikoYamashita,anHHMIinvestigatorattheUniversityofMichigan.“Tomakesureyourdaughtercanmakeadaughterwhocanmakeadaughter,ithastobeperfect.”

Toachievethatperfection,GSCshaveevolvedsophisticatedstrategiesfortheirpreservationandfunction.Bystudyingthosestrategies,researchershaveaccomplishedmanyfirstsinstemcellbiology,includingidentifyingthefirststemcellniches,discoveringnovelmechanismsofcelldivision,andpinpointingkeysignalsthatregulatestemcellactions.Andmanyofthemolecularpathwaysgoverningtheprecisebalancebetweenself-renewalanddifferentiationhavebeenillustratedinvividdetail,makingGSCsarguablythebestunderstoodstemcellsinbiology.

Walking the LineGermlinestemcellsareawalkingcontradiction:theyareultimatelyabletogenerateeverycellinthebodybutmustproduceonlyonetypeofcell—spermoregg.“Thegermcellsareveryprimitiveembryoniccells,andit’sreallyimportantforthemtoavoidsomaticdifferentiation.Theyshouldn’tbecomemuscle.Theyshouldn’tbecomenerve,”saysLehmann,whostudiestheveryearlieststagesofGSCformationinfruitflies.

Fromtheverybeginning,theprecursorstoGSCs,knownasprimordialgermcells(PGCs),areprotectedfrombecomingsomaticcellsbyformingattheedgeofanembryo,awayfromsignalsthatmightinducethewrongfate.Thisoccursbyoneoftwomechanisms.Inmiceandothermammals,signalspassedbetweenextra-embryonictissuesandtheembryocoaxaspecificsetofcellsattheperipherytobecomethegermcells.Inflies,worms,frogs,andfish,ontheotherhand,aspecialprotein-richzoneinthecytoplasmoftheegg,calledthegermplasm,determineswhichcellsbecomePGCs.GermplasmisathicksoupofproteinsandRNA,withamuchhigherconcentrationofmoleculesthanintherestofthecytoplasm.

In1992,attheMassachusettsInstituteofTechnology(MIT),Lehmannidentifiedakeyingredientinthatsoup—amaternaleffectgeneshenamedoskar,whichdepositsacriticalproteinintothegermplasm.InDrosophila—thosesmall,hairyfruitfliesoccupyinglaboratoryvialssince1901andafavoriteforstudyingdevelopment—ifoskarisinactive,nogermplasmforms,andnogermcellsdevelop.

Butwhenoskarisactive,PGCsdoform.Fromthere,theyadoptacleverdefensivemaneuvertoavoidrapidlydividingandspecializingliketherestofthecellsintheembryo.ThePGCstakeabreak;theypause.

Thishappensthroughextremetranscriptionalrepression,whereinDNAispreventedfrombeingtranscribedintoRNAandisheldtemporarilysilent,asifinatrance.Variousmolecularmechanismscontrolthisrepression,dependingonthespecies,buttheoutcomeisthesame:littletonotranscriptionandthereforenoexpressionofthegenome.

Oneofthosemechanisms,recentlydiscoveredbyHHMIInvestigatorBradCairnsattheUniversityofUtah,isthe L

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the earliest stages of germline stem cell formation in fruit flies.


Brad Cairns’ team identified a slew of chemical tags that activate or silence genes in germ cells.

Yukiko Yamashita is focused on why one stem cell stays in the niche while another moves away.

modificationofthegenomebychemicaltagsthatbindandsilencedevelopmentalgenes,thuspreventingdifferentiation.Inaprojectthattookmorethanfourandahalfyears,Cairns’teamlookedforandidentifiednumerousmodificationsofDNAandhistones—theproteinsthatpackageDNA—inmultiplestagesofmalegermcelldevelopmentinmice.Theyalsovalidatedtheirfindingsinhumansperm.

Overtime,theprojectbecamebiggerandbigger.“Wewereastoundedtofindsomanychemicalmarksandsomanychangesatdifferentdevelopmentalstages,”saysCairns.“Wemadeamajoreffortforseveralyears,aseachnewresulthelpedexplainthelogicoftheregulationprocess.”AstheyreportedinAugust2014inCell Stem Cell,thegroupfoundaslewoftags—half“activating”tagsandhalf“silencing”—stucktokeydevelopmentalgenes.Theactivatingtagshelpensurethatthegenescanbeturnedonafterfertilization.Untilthattime,thegenesareheldincheckbythesilencingtags.

WiththeDNAofgenespromotingdevelopmentinhibernation,someothersystemmustregulatetheactivityofPGCs,whichincludesmovingtothegonadandbeginningtodivideandproducedaughtercells.Itturnsoutthatahallmarkofearlygermcellsissmall,stickyballsofRNAandproteinsthatmovearoundthecytoplasmandregulateRNA,leavingtheDNAalone.ResearchersbelievethattheDNAinsideGSCsissoprecious—theinstructionsforthesurvivalofthespecies—thattheDNAiskeptinapristinestateinthenucleus,whiletheRNAishighlyregulatedbythesestickyRNA-proteinclumpsinthecytoplasm.“HavinglessofanemphasisontranscriptionandmoreonRNAregulationmayhavesomethingtodowithpreventingdamagetotheDNA,”saysLehmann.“Wedon’treallyknow,butitisaninterestingphenomenon.”

InDrosophila,PumilioandNanos—proteinsLehmanndiscoveredasagraduatestudent—aretwooftheproteinsthatcomposetheseRNA-proteinclumps,andtheycanbefoundingermcellsfromwormstohumans.InthewormCaenorhabditis elegans,JudithKimble,anHHMIinvestigatorattheUniversityofWisconsin-Madison,identifiedaPumilio-likeproteincalledFBF,requiredforGSCmaintenance,whichrepressesmorethanathousandRNAsingermlinestemcells,manyofwhichactivatedevelopmentalpathways.

A Niche RoleKimble’sworkonGSCsstartedmuchearlier.In1981,asapostdocattheMRCLaboratoryofMolecularBiologyinCambridge,England,sheusedalasertodestroycellsinthetipofthedevelopinggonadofC. elegans.Bydoingso,sheidentifiedasinglecell—dubbedthedistaltipcell—thatwasabsolutelyrequiredformaintainingtheworm’sGSCs.Thisspecialregulatorycellwasnotagermcellbutasomaticcell,andwhenKimblemoveditaroundthegonad,theGSCsmovedwithit.ShewasthefirsttoshowthatanearbysomaticcellisessentialforGSCmaintenance—suggestiveofastemcellnicheinC. elegans.

Subsequently,KimbledemonstratedthatthedistaltipcellnichemaintainsstemcellsbyproducingamoleculethatbindstoaNotchreceptorproteinonthesurfaceofGSCs.ItwasthefirsttimethatNotchsignalingwasshowntocontrolstemcells.ResearcherswouldgoontodiscoverthatNotchisapowerfulregulatorofmanytypesofstemcellsandanimportantdriverofcancer.

KimbleandherteamhavesincefocusedonunravelingtheelaboratenetworkdownstreamofNotchsignaling,aseriesofabout100moleculesthattogethercontrolthebalancebetweenself-renewalanddifferentiation.Byidentifyingandmanipulatingthosemolecules,herteamhasbeenabletoexpandandcontractthepoolofGSCsin C. elegans.

Kimblehasnoplanstostopuntilshe’sidentifiedallthemajorlinksinthestemcellNotchsignalingpathway.

“They’re the only cells that you don’t actually need for your own survival, but you do need them for the survival of your species.”—ruthlehmann

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“Iwantthewholenetworkthatregulatesthisdecisionbetweenself-renewalanddifferentiation,”shesays.Andshe’swellonherway:justlastspring,intheMarch11,2014,issueofProceedings of the National Academy of Sciences,KimblereportedtwonewproteinsessentialforGSCmaintenancethataredirecttargetsofNotchsignalinginGSCs—alongsoughtmissinglinkbetweenNotchregulationandthedownstreamregulatorynetwork.

Bythe1990s,mostresearchersbelievedintheexistenceofastemcellniche—aspecializedlocationinsideatissuewherestemcellsreside—buthadneverdirectlyobservedindividualstemcellsinsideaniche.Atthetime,itwasclosetoimpossibletoreliablyisolateasinglehuman—ormouse—stemcell,muchlesstoidentifywhereinthebodythecelloriginated.

However,inDrosophila,theproblemwastractable.“Thesmallsizeandsimplicityoftheirtissuesmakeiteasytolookatwhatindividualcellsaredoing,”saysAllanSpradling,anHHMIinvestigatorattheCarnegie

InstitutionofWashington.“Nowhereinallofbiology,thatweknowof,isthismoretruethanwiththestemcellsoftheDrosophilaovary.”

Eachfruitflyovarycontains12to16flexibletubesfilledwithchainsofmaturingeggcells.Tolocatethestemcellsintheovary,onesimplyfollowsasingletubebacktoitsstart,totheverytipwherealltheothercellsoriginate.Thereresidetwoplump,

roundgermlinestemcells,thecellsthatproduceeggs.JusthowmanyanimalspecieshavefemaleGSCsisstilluncertain,thoughevidencesuggeststhatfemalemammals,includinghumans,arebornwithalltheeggstheyareevergoingtoneed.

ClassicalhistologistsidentifiedDrosophilagermlinestemcellssimplybylookingunderamicroscope,butSpradlingarguedthattheidentityofsuchcellscouldbeknownwithcertaintyonlybytracingtheircelllineages—thatis,byidentifyingalltheprogenyofasinglecell.In1995,heverifiedtheconclusionsoftheclassicalhistologistsaboutDrosophilaovarianGSCsbydoingso,butinmanyothercases,suchanalysisshowedthatearlyguessesaboutothertypesofstemcellswerenotcorrect.

In2000,heandpostdocTingXiealsousedlineagetracingtoidentifyagermlinestemcellnicheinDrosophila.TheymarkedovarianGSCswithageneencodingafluorescentprotein,makingthecellsglowundercontrolledconditions,andthenmanipulatedthem.Whentheresearchersremovedoneofthetwocells,theremainingcellreplicated,anditsoffspringtooktheplaceoftheabsentstemcell.SpradlingandXierealizedthatthelocation,notsomemagicalaspectofthecellitself,waskeytotheexistenceofastemcell.“Ourworkshowedastemcellisreallynotaspecialcell;itisjustanaverage,undifferentiatedgermcell,”saysSpradling.“Thelocationmadeallthedifference.”

Spradlingthenturnedtothegenesthatmightbeinvolvedinself-renewalanddifferentiationintheDrosophila ovary.Hefoundonesterilemutationthatcausedthetipofeachtubeintheovarytofillupwithstemcells,likeabagofmarbles.Namingthegenebag of marbles,orbam,Spradling’steamconcludedthatthegeneisrequiredfordifferentiation.Next,Spradling’slab,incollaborationwiththeUniversityofTexasSouthwesternMedicalCenterlabofDennisMcKearin,nowaseniorscientificofficeratHHMI,identifiedasignalingpathwayinthenichethatenablesGSCstosilencebamandretaintheirstemcellidentity.ButwhenGSCsdivideanddaughtercellsmoveoutsidetheniche,theynolongerreceivetheproteinsignal,so bam isactiveandthecellsdifferentiate.

Orientation MattersAroundthetimeSpradlingwasidentifyingthesignalsregulatingthefemaleDrosophila niche,Margaret“Minx”Fuller,atStanfordUniversity,wassimultaneouslystudyingGSCsintheDrosophila testis.Unlikethefruitflyovary,thetestisnichecontainssixto12germlinestemcellsclusteredaroundagroupofsomaticsupportcells,calledthehub,likepetalsonaflower.YukikoYamashita,thenapostdocin

Judith Kimble was the first to show a somatic cell is essential for GSC maintenance.

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“Our work showed a stem cell is really not a special cell; it is just an average, undifferentiated germ cell. The location made all the difference.”—allanspradling


Fuller’slab,decidedtotakeaquicklookatmitosis—theprocessofcellulardivision—inGSCs.YamashitaandFullerhopedthatbystudyinghowGSCsdivide,theymightgaininsightintohowoneoftheoffspringremainsastemcellwhiletheotherdifferentiates.

“Webothplannedjusttohaveatinypaperoutofthatproject,”saysYamashitawithalaugh.“That’snotwhathappened.”Yamashitafoundthatthecentrosomes,small,anchor-likeorganelleswherethestructuralfilamentsneededforcelldivisionoriginate,werenotbehavingasexpected.Mostbiologytextbookshavediagramsshowingcentrosomesseparatingafteracellcommitstomitosis.YetinthemaleflyGSCs,centrosomesweresplittingapartfromeachotherandmovingaroundthecellbeforetheinitiationofmitosis.Becauseofthisearlymovement,thecentrosomeshadmarkedwherethemitoticspindleshouldbeoriented—perpendiculartothestemcellniche,sothatonecellstaysinthenicheandtheotherispushedout—beforemitosisevenbegan.

“That[orientation]isprobablyresponsibleforthecontinuedbalancemaintainingthestemcellpopulationandcontinuingtoproduce[sperm],”saysFuller.“Itisreallycriticalforthestemcellprogram.”Inretrospect,saysYamashita,itmakessensethatgermlinestemcellswouldtakethisextra,earlyprecautiontoprepareforreliablestemcelldivision.Their“tinypaper”waspublishedin2003inScience andhassincebeencitedover415times.

YamashitahascontinuedtoexplorehowonedaughtercellstaysinthenichetobeaGSCwhiletheothermovesaway,tocommittobecomingaspecializedspermcell.“Whatkindofprecisemechanismcanmakesurethatonlyonecelloftwodaughtercells,nexttoeachother,cheektocheek,getsasignaltomakeitastemcell?”Yamashitaasks.“It’slikeyouhavetwintoddlersandtrytogiveasnacktoonlyoneofthem!”

Recently,oneofYamashita’spostdocscametoherwithamicroscopeslideandaskedhertolookclosely.There,inDrosophilaGSCsinthetestis,sheobservedaslimwhiskerofmicrotubules,anano-sizedbridgepokingoutofthestemcelltotouchthenearbystemcellniche.Yamashitawassosurprisedshewentbacktotissueimagescollectedsome10yearsearlierthatwerestoredonhercomputer.Loandbehold,therewasthesamewhisker.“Itwasthere;Ijusthadn’tnoticedit,”shesays.“Wemissedthatstructureforsuchalongtime.”TheteaminvestigatedanddiscoveredthatacriticalsignalissentfromthenichecellstotheGSConthesurfaceofthatwhisker—aprecisemechanismforsignalingtothestemcellbutnotthedaughtercell.

Model BehaviorOnceadaughtercelldoesleavetheniche,itbeginstheprocessofbecomingaviablegamete,whichstartswithmeiosis—theprocessofcelldivisionthatleadstogametes,eachwithonlyhalfthegeneticinformationneededtoformanindividual.

AttheWhiteheadInstituteandMIT,DavidPageandcolleagueshavedecipheredthesignalsthatinitiatemeiosisinbothmaleandfemalevertebrates,includingthepowerfulStra8(Stimulated by retinoic acid gene 8),ageneexpressedinthefetalovaryandjuveniletestis(mammalscontinueproducingspermintoadulthood,thoughnoteggs),rightatthetimemeiosisisinitiated.“Adefiningfeatureofgermcellsistheyallknowhowtodomeiosis,”saysPage,“soIwantedtoknowhowthissuper-conservedprogramisinitiated.”

Pageandhisteamhaveshownthatretinoicacid,afleetingandhard-to-trackchemicalbelievedtobeproducedbynearbysupportcellsinthegonad,activatesStra8inbothovariesandtestes.InAugust2014,Page’steamreportedinPLoSGeneticsthatretinoicacidactivatesa secondgene,Rec8,whichencodesaproteinalsocriticalformeiosisandisconservedallthewayfromyeasttohumans.

“Thereissomuchnowknownfromtheflyandwormmaleandfemalegermlinesabout[numerous]thingshappeninginstemcells,”addsFuller.“Thesehaveprovidedparadigmsforpeopleworkinginotheradultstemcells—modelsforwhatkindsofthingstolookfor.”

Allan Spradling’s team was able to identify a stem cell niche in Drosophila.

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In Drosophila testes, germline stem cells (violet) cluster around a group of support cells called the hub (bright red).

Power to the PosseServing as both

a lifeline and a launching pad, the Posse STEM

program promotes persistence among

multicultural college students.

Power to the PosseServing as both

a lifeline and a launching pad, the Posse STEM

program promotes persistence among

multicultural college students.

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Fforbrynmawrcollege sophomoreFransheskaClara,goingtocollegewasn’tjustapersonalgoal:itwasafamilyone.Neitherherfather,whohailsfromElSalvador,norhermother,anativeofPuertoRico,hadgonetocollege.AndtheyhadcometoAmerica—toWorcester,Massachusetts,anhouroutsideBoston—inthehopethattheirtwokidscouldgetacollegeeducationandfulfilladreamthattheyhadn’tbeenabletoachievethemselves.

Clarawasdriventosucceedinhighschool.Shegotgoodgrades.Shewaspresidentofherschool’sSTEMclub,agroupfocusedonscience,technology,engineering,andmathematics.AndshespentasummerandafullsemesterdoinginternshipsthroughthebiologyandbiotechnologydepartmentsofWorcesterPolytechnicInstitute.

SowhenshefirstheardaboutthePosseSTEMprogram—whichwouldsend10-person“Posses”oftheverybeststudentstotop-notchcollegesacrossthecountrytofocusonscience—Clarawaseagertolearnmore.Theprogramisdesignedtoidentifyhigh-achievingstudentleadersinurbanhighschoolsandsendthemtocollegeasagroupsotheycanleanononeanotherforsupport.ThePossemembersarealsopairedwithmentorsatthecollegesandreceivesignificantaccesstoscienceresearchopportunities.Itisaprogramtailoredtohelpthesestudents,whooftencomefromdisadvantagedbackgrounds,succeed.

AteachernominatedClarafortheprogram,andshespentmonthsgoingthroughdemandingleadershipexercises,interviews,and

problem-solvingprojectsalongsidesome100othersimilarlytalentedstudents.“Everyonewassoimpressive,”sherecalls.“IalwaysfeltlikeIwouldn’tgetthroughtothenextround.”

ButonDecember3,2012—threedaysbeforeher18thbirthday—afterafinal,exhaustingthree-hourinterview,shegotthenews:shehadbeenselectedforBrynMawr’sfirstSTEMPosse.“Ireallyrememberthatcall,”shesays.“Mydadwascrying.Mymomwascrying.Itwasareallybigthingforus.”

ForstudentslikeClara,thePosseSTEMprogramrepresentsbothalifelineandalaunchingpad.Thefour-year,full-tuitionscholarshiphelpsstudentswhosefamiliesmightnototherwisebeabletoaffordacollegeeducation.Andthewide-rangingsupportnetworkoffaculty,staff,andnineotherhigh-achievingstudents,allaimingatsimilargoals,helpsstudentsfocusonachievingtheirdreams,oftenfasterthantheyeverthoughtpossible.

WiththehelpofBrandeisUniversitychemistryprofessorIrvEpstein,whowassupportedwithagrantasanHHMIprofessor,thefirstSTEMPossearrivedatBrandeisin2008.Sincethen,nineotherschoolshaveadoptedtheprogram,andeachwillsupport50newSTEMPossestudentsoverthenextfiveyears—a$70-millioninvestmentin500studentswhowillhelpshapethefutureofscience.Theinvestmentissignificant.Butthelong-termpayoff—forstudents’individualcareersandfordiversityinthesciences—couldbeevenbigger.

Model of SuccessTheoverarchingPosseprogram,thebrainchildoffounderDeborahBial,hasbeenaroundsince1989.Bialwasinspiredtostarttheinitiativewhensheheardastudentwhohaddroppedoutofcollegesay,“IneverwouldhavedroppedoutifI’dhadmypossewithme.”Sheknewthatkidsfrombig—andoftenunderfunded—urbanpublicschoolsystemsoftenstruggledandthendroppedoutofelitecolleges,eventhoughtheywerejustastalentedastheirpeers.ThePosseprogramaimedtoreducethesedropoutratesbyrecruitinggroupsofstudentsfromspecificcitiesaroundthecountrywhowouldattendthesamecollege,supportoneanotheralongthejourney,andhelpmakethecampusmorewelcomingtostudentsfromallbackgrounds.

Eachstudentreceivedafour-yearscholarship,on-campusmentors,andthecamaraderieoffellowPossememberswhohadallgraduatedfromhigh

“We know there are extraordinary numbers of talented students who fail to attend colleges that fit with their high school achievements.”—danporterfield

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schoolsinthesamecity.“IfyougrewupintheBronx,forexample,butyouendedupinMiddlebury,Vermont,you’dbealittlelesslikelytoturnaroundandgohomeifyoufeltcultureshock,”explainsBial.Theresultshavebeennothingshortofexceptional:therearemorethan2,600alumnioftheprogram,andthegraduationrateforPosseScholarsis90percent.Theprogramdoesnotexplicitlyseekstudentswhohavehighneedorwhoareminorities.However,becauseofthedemographicsoftheschoolstheprogramdrawsfrom,80percentofPosseScholarscomefromeconomicallydisadvantagedbackgroundsandareAfricanAmericanorLatino.Theprogramisextraordinarilycompetitive:thisacademicyearalone,thePosseprogramreceivedmorethan15,000nominationsfor700slots.

Brandeis’Epsteinhasseenthevalueoftheprogramfirsthand.HewasinvolvedinBrandeis’traditionalPosseprogramwhenhewastheschool’sprovost,andheknewstudentsthrivedinit.

Whenhesteppeddownasprovostandreturnedtoteaching,hebecamefrustratedbythelackofdiversityinhisscienceclassrooms—andconcernedthatsomeofthedynamicsofthePosseprogrammightbepushingsmartkidsawayfromthesciences.“Whena[Possestudent]gota40onherfirstchemistrytest,shemightsay,‘Ican’tdoit,’andtheotherPossestudentsmightsay‘You’resmarterthanweare,andwe’redoingjustfineinourclasses;maybeyoushouldswitchoutofthatclass,’”Epsteinsays.

Thesolution,hebelieved,wasn’ttoputmorepressureonthoseindividualstudentspursuingthesciences—itwastocreatePossesinwhichsciencewasthefocus.“Ithoughtiftheywerealldoingscienceandallcommittedtoit,thenthepressurewouldbeinward,towardscience,ratherthanoutward,awayfromit,”hesays.WorkingtogetherwithBialandBrandeisadministrators,andwiththesupportofhisHHMIprofessorship,EpsteinhelpedbringthefirstSTEMPossetoBrandeis’campusin2008.

ManycomponentsofthePosseSTEMprogramatBrandeiswerenearlyidenticaltothoseofthetraditionalPossemodels:arigorous,months-longrecruitingprocesstoidentifygreatstudentswhowerepotentialleaders;pre-collegemeetingsforstudentstogettoknowotherPossemembers;andweeklymeetingswithstaffandfacultymentorsandtheirPosseduringtheschoolyear.ButthePosseSTEMmodellayeredontwoadditionalpieces.Onewasatwo-weekimmersionprogrambeforestudents’freshmanyearthatintroducedthemtotheircampus,toscienceatanelitecollege,andtoimportantsupportservicessuchastutoringcenters.ThesecondwasamentorforPosseSTEMstudentswhohadabackgroundinthesciences.

InitialresultsfromtheBrandeisprogramhavebeenpromising:29ofthe30studentsinthefirstthreePosseshavegraduated.Ofthose,22earneddegreesinthe“hardsciences”ofbiology,chemistry,physics,

math,orneuroscience;theothersevenearneddegreesinpsychologyorsciencepolicy.

EncouragedbyBrandeis’pioneeringwork,fourotherschools—theUniversityofWisconsin–Madison,BrynMawrCollege,Franklin&MarshallCollege(F&M),andTexasA&MUniversity—adoptedthePosseSTEMprogrambetween2008and2013.AccordingtoF&MPresidentDanPorterfield,hisschool’sinvestmentintheprogrammakesperfectsense.“Weknowthereareextraordinarynumbersoftalentedstudentswhofailtoattendcollegesthatfitwiththeirhighschoolachievements,”hesays.“Weinvestinprograms,includingPosse,toprovidestudentsaccesstohighereducationthat’scommensuratewiththeirtalent.Andwebelievetheywillgoontomakeadisproportionatedifferenceintheworld.”

In2015,withencouragementfromPresidentObamaataWhiteHouseSummitonhighereducation,fivemoreschools—DavidsonCollege,GeorgetownUniversity,SmithCollege,MiddleburyCollege,andPomonaCollege—willbeginmakingthatinvestmentbyaddingSTEMPossestotheirstudentbodies.Bialbelievestheimpactcouldbetransformative.“Wehopethatwe’llseesimilarsuccessratesatalloftheseschools,”shesays.“Itwillprovetothenationthatthere’sapoolofyoungpeopleouttherewhoarenotonlycapableofbeing,butwhowillbe,superstars.AndtheywillhelptheseinstitutionsbringdiversityintotheirSTEMmajorsinawaythathasn’thappenedbefore.”

Practical, and Moral, SupportSTEMPossesaredesignedtohelpstudentswithwhateverproblemsthey’refacing.

FransheskaClara,forexample,sayssheleanedonherPosseforsupportwhenshemissedherfamilyandfriendsfromhome.“Theothers

As members of the Bryn Mawr STEM Posse, Fransheska Clara (left) and Carol Bowe help each other navigate social adjustments as well as academics.

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werealsostrugglingwithhomesicknessandtransitioningintoanewenvironment,”shesays.“Itreallyhelpedtohavethemaroundandtotalkaboutthings.Irelatedtothem.Iknewtheyfelttheexactsameway.”

ForUniversityofWisconsin-MadisonsophomoreCarolineRozado,herPossewastheretohelppreventsmallproblemsfromspiralingintobigones.RozadohadbeenanaccomplishedstudentatherNewYorkCityhighschool,andherextracurricularactivitiesincludedthreeyearsofinternshipsfocusedonecogeneticandenvironmentalresearch.Butevenso,shewasworriedaboutherintroductorychemistryclassatWisconsin.“Myhighschoolhadlowfunding,sotheycouldn’tfindachemistryteacher,”shesays.“Instead,theyhadabiologyteacherteachchemistry,butevenourbiologyteachercouldn’tgetustheinformationweneeded.”

Itwasn’tjustthecoursematerialatWisconsinthatseemeddaunting:she’dneverbeeninaclassroomwith300studentsbefore.“Itwasnervewracking,”shesays.“WasIsupposedtositinthefront?Themiddle?Theback?”

Thefirstcoupleofweeksofherchemistryclassweretough.ButherPosse—andhermentor,EmilieHofacker—hadherback.Hofacker,whoisassistantdirectorforSTEMinitiativesatWisconsin,gaveherapeptalk,andanolderPossestudentgavehertipsonhowtoreadthetextbookandwhatproblemstofocuson.Soon,sheandaPossememberfromhercohortworkeduptheircouragetogotogethertoofficehourswiththecourse’sprofessor.Bytheendofthefirstmonth,Rozadowasontrack.

Thehelpsheneededwassimple,buttheimpacthasbeenprofound.“I’mnotafraidtoaskprofessorsquestionsanymore,eveniftheyseemreallybasic,”shesays.“Professorsactuallywanttoknowwhatwedon’tunderstand,andIjustneededtohavethosefirstexperiencestogettothepointwhereIfeltitwasokaytotalktothem.”

ForHofacker,helpingstudentshaveepiphanieslikethesearecentraltoherwork.“Sometimes,studentsjustwanttoknowthatit’sgoingtobeokay,”shesays.“Therewillbedayswhentheywanttogiveup,butwe’llhelpthemworkthroughit.”

STEMPossesarehelpfultonavigatesomeofthesubtleraspectsofthesocialadjustmentaswell.BrynMawrsophomoreCarolBowesaysthatdespiteanoverwhelminglypositiveexperienceincollege,shehashadtodealwithstudentswhomakecondescendingremarkstoherbecauseshe’sinaSTEMPosse.“Sometimesthere’sastigma,”shesays.“There’sanideaof,‘You’reonascholarship,andhadyounotbeenonascholarship,youwouldn’tbehere.’Peoplesaysomanythingsaboutmoney.”

ShesaysherPossediscussedhowtohandlesuchissuesduringtheirfirstmeetingsinBoston,beforetheyarrivedoncampus.“It’seasytosay,‘Oh,I’lljusteducatethem,’”Bowesays.“Butwhenyou’restandinginfrontofapersonwhosaidsomethingignorantandyoufeelreallyhurt,youmightnotactuallyknowhowtodothat.”ShesaysshewasrelievedtobeabletotalktootherPossememberstofigureouthowtonavigatethenuancesofsuchconversations.

AsPosseSTEMstudentsfocusontheirsciencestudies,theyalsobenefitfromtwootherPosseperks:researchopportunitiesandlabtime.PosseSTEMstudentsoftenhearaboutresearchopportunitiesbeforeotherstudentsoncampus,andsomeon-campusresearchersspecificallyrequesttoworkwithPosseSTEMstudents.Evenifstudentsdon’tpursueresearchfromtheoutset,theystilloftenrubshoulderswithresearchersincampusjobsscrubbingbeakersinalabinsteadofpotsandpansinthecafeteria.

Forstudentscomingfromdisadvantagedbackgrounds,thattinydistinctioncanbecritical.“Somanystudentshaveanotionthatscienceissomethingwherealonescientistworksinthedeadofnightwithnohumancontact,”saysBrandeis’Epstein.“Butthesekindsofworkexperienceshelpthemseethatresearchgroupsaretight-knit,multi-generational,multi-ethniccommunitiesthatsupporteachother.Ifyouwashdishesinalab,eventuallyyou’llprobablyendupdoingresearchinalab.”

SuchwasthecaseforRozado,whowasabletoparlayherexperiencecleaningglasswareintheschool’sresearchdemonstrationlabintoworkwithgeneticsprofessorJerryYin.She’shelpingYinwitharesearchprojectstudyingproteinsignalingandproteinlevelsinpeoplewho

When University of Wisconsin sophomore Caroline Rozado (left) needs support, she can turn to her mentor, Emilie Hofacker.

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haveneurodegenerativediseases.“It’ssogreattoknowthatresearchersareopentostudentsworkingintheirlabwiththem.They’repatientenoughtoteachustheseprotocols,andevenifwemakemistakes,they’llhelpusfixthem,”shesays.“It’samazing.Itfeelslikeanhonor.”

Metrics of SuccessThefourmostrecentPosseSTEMprogramsaretoonewtohavesignificantdatacomeoutofthem,buttheschools’administratorsarealreadythrilled.F&M’sPorterfield,forexample,saysthatfromthethreePosseSTEMgroupscurrentlyoncampus,notoneofthestudentshasdroppedout.Theoldestcohort,nowjuniors,hasanaverageGPAthatishigherthantheirnon-PosseSTEMclassmatesinascience-heavycurriculum.Meanwhile,BrynMawrreportsthatallbutonememberofitsfirstSTEMPossehavecontinuedontotheirsecondyear,withthe10thlikelytoreturnforthespringsemester.Wisconsin’sHofackerreportsthatnotonlyareherschool’sPosseSTEMstudentsperseveringinthesciences,buthalfareontracktograduatewithSTEMmajorsinfouryears,eventhoughthetypicaltrajectoryattheschoolforsuchmajorscandemandanextrasemesterortwo.

Epsteinaddsthat,insomeways,thesestudentsaredefyingexpectations.TheaveragemathandverbalSATscoreforanincomingBrandeisstudentis1353;foraPosseSTEMstudentit’sinthemid-1100s.Yetdespitethat200-pointgap,STEMPossestudents,sofar,haveafour-yeargraduationrateof97percent,comparedtotheschool’soverallaverageof86percent.ToEpstein,it’sanindicatorthatthePosseSTEMprogramisfocusingontherightthings.“Ithinkwe’vepickedpeoplewhoseabilitiesarehigherthantheirtestscoresalonemightsuggest,”heexplains.Withtherightsupport,they’renotjustpersistinginschool—they’resucceedingatthehighestlevels.

STEMPossestudentsaren’ttheonlyonesbenefitingfromtheprogram:facultydiscussionsaboutthePosseSTEMprogramandteachingsciencemoreeffectivelyhavespilledoverinwaysthatbenefitallstudents.AtBrynMawr,forexample,thePosseSTEMprogramopenedupalargerconversationattheschoolaboutsupportingstudentsintheSTEMfields;thecollegeiscurrentlypilotingacoursecalled“FundamentalsofMathematicsforScienceandSocialScienceStudents,”whichgivesstudentsthequantitativeskillsthey’llneedtosucceedinintroductorysciencecourses.AtF&M,administratorshavebeensopleasedwiththecohort-mentoringmodelthatthey’veexpandedittosixgroupsoffirst-generationcollegestudentswhomeetregularlywithfacultymentorstobolstertheirchancesofsuccess.

AndF&MchemistryprofessorKenHess,whoservedasamentortothefirstgroupofPosseSTEMstudentswhentheyarrivedoncampus

in2012,sayshisexperiencewiththePossestudentshashelpedhimbeabettermentortoallofhisstudents.“I’mmuchmoreholisticinthewayIthinkaboutthingsnow,”hesays.“I’mfarmorepatient.I’mabetterlistener.I’mmoresensitiveto,andappreciativeof,thechallengesthatstudentsbringtotheclassroomandhowthatmightaffecttheirperformance.”

A Path ForwardBecausethePosseSTEMmodelsocloselyfollowsthewildlysuccessfultraditionalPossemodel,andbecauseearlyPosseSTEMoutcomesdataseemsopromising,it’snotdifficulttoimaginethatitslong-termsuccesswillbesimilar.AlumnifromtraditionalPossecohortshaveearned43Fulbrightscholarshipssince2007.And41percentofPossealumniwhoaretwoormoreyearsoutfromgraduatinghaveearnedagraduatedegreeorarepursuingagraduatedegree.

PosseSTEMstudentsarejustasambitious:CarolineRozado,whoplanstopursuemajorsinneurobiologyandcommunications,saysherdreamistobethenextDr.Oz,someonewhousesherdeepknowledgeofsciencetoteachtheworldhowourbodieswork.ShecreditsthePosseprogramforgivingherthechancetotalkwithotherbrightsciencestudentswhohelpedherunlockherpassion—andwhofuelherrelentlessdrivetosucceed.“ThoseconversationsmademerealizehowhappyIamwhenIcanhelpotherslearn,butIdidn’tknowthatuntilIwasinPosse.NowIunderstandwhyI’mdoingthisworkinscience.”

ForFransheskaClara,Possehasgivenhertheonepieceofthepuzzleshewaspreviouslymissing:confidence.“Iusedtobeverydoubtfulanduncertain,butnow,IknowIcanmatchupwithanyone,”shesays.“That’shelpedmewithacademics.It’shelpedmewitheverything.”

“Sometimes, students just want to know that it’s going to be okay.There will be days when they want to give up, but we’ll help them work through it.”—emiliehofacker

Perspectives & Opinions


Jorge Dubcovsky—Jorge Dubcovsky heads the Wheat Breeding Program and Wheat Molecular Genetics Laboratory at the University of California, Davis.

Rethinking GMO Regulation Areweoverregulatingtransgeniccrops?In the United States, as in most countries, genetically modified crops are heavily regulated at every stage of their development—from research through marketing. While oversight is valuable, too much can be detrimental, believes HHMI-GBMF Investigator Jorge Dubcovsky. He explains why our current approach to transgenic regulation may not be in society’s best interest.

Why are people afraid of GMOs? Peopledon’tknowwhatGMOs,geneticallymodifiedorganisms,are.Theyjustknowsomebodyismessingwiththeirfoodandworrywhetherthatpersonhasgoodorbadintentions.Thoughfearissometimesirrational,it’svalidtofearwhatyoudon’tknow—weneedtorespectthatfeeling.

Unfortunately,governmentsoftenrespondtosuchfearbyincreasingregulation.Inthecaseofwheat,forexample,thetestingandregulatoryprocessissoburdensomeit’salmostimpossibleforpublicbreedingprograms—atland-grantuniversitiesortheDepartmentofAgriculture—toreleasetransgenics.It’sokaytoregulateGMOs,butcurrentlyweregulatethetechnology,nottheproducts.Weshouldlookatwhatwe’reputtingintheplant,nothowit’smade.

What’s wrong with regulating the technology? It’sirrational.IfIaddanewwheatgenetoawheatseedtoincreaseitsresistancetodiseaseoritsnutritionalvalue,

theresultingplantisconsideredtransgenic.EvenifithasnoforeignDNA,thenewvarietycanbereleasedonlyafterlengthyandexpensivetests.ButifIachievethesameresultbyrandommutagenesisorbyrecombinationofawheatchromosomewitharelatedwildspecies,it’snotconsideredaGMO,andIdon’tpayacent.

Usingthesameprocess,youcanmakeharmlessproducts,likemorenutritiousvarietiesofwheat,orproductswithhigherrisk,likeplantsthatexpresspartofavirustoconferresistanceagainstpathogens.Idon’tseethelogic,scientificallyspeaking,ofregulatingtheprocessratherthantheproduct.

What are the consequences of regulating the process? It’snowsodifficulttodeveloptransgeniccropsandsoexpensivetoreleasethemthatpublicbreedingprogramscan’taffordtodoso.Thisposesariskthatlargecorporationscouldcontroltheseedmarket—andtheymaymakedecisionsbasedonthebottomlineratherthanpublicbenefit.

Fortunately,we’renotinthatsituationyet,atleastforwheat.In2012,roughly68percent

ofthewheatplantedintheU.S.representedvarietiesproducedbythepublicsector.Butinthefuture,ifcurrentregulationsremaininplace,thepublicwheat-breedingsectorwon’tbeabletocompete.Overregulationcouldleadtobigcompaniescontrollingtheseedsupply.Keepingpublic-sectorinvolvementoffersseveraladvantages.Publicbreedingprogramshelptrainnewplantbreeders;theyserveasabackupfor

industryefforts;andtheycanfocusonlonger-termgoals,likeincreasingcropdiversity,andontraitsnoteconomicallyattractivetocorporations.

How can we improve things? Ibelieveahybridmodel,withbothpublic-andprivate-sectorbreedingprograms,offerstherightbalance.Thepublicsectorhasfreedomtoexploretraitsbeneficialtoconsumers,

whilecompaniesaregenerallybetteratdistributingandsellingseedsandcanbringadditionalresourcestobreedingefforts.WehaveasystemlikethatinCalifornia,anditworkswell.Companiesgetnewtechnologyandgermplasmfrompublicbreedingprograms,growersgetthebestseeds,andsocietygetsimprovedproductsandreduceduseoffungicidesandpesticides.

Ialsobelievepublicbreederscanhelpeasefearoftransgeniccrops.Forexample,wecancreatetransgenicsbyintroducingDNAonlyfromthesameorrelatedspecies(processesknownas“cisgenics”and“intragenics”).OrwecanuseapproachesknownasRNAinterferenceandCRISPR,whichjustknockoutcertaingenes.SuchapplicationseliminateintroductionofforeignDNA,somayreducethefear,butthey’restillverypowerful.Forexample,RNAinterferencecanbeusedtoreduceglutenintoleranceandincreasethefiberinagrain.Ifregulationsarereducedfortheselesscontroversialtechnologies,publicresearcherscanusethemtogenerateusefulproducts.

Are things likely to change? Yes,people’sfearwilllikelydiminish.Thereweresimilarconcernsmanyyearsagowhengrowersintroducedhybridcrops.Today,nobodycaresabouthybrids.Geneticengineeringisapowerfultechnology.Itletsusdolotsofpositivethingsinafaster,moreefficient,verycontrolledway.Andit’shappening.Ifyoulookatmaizeandsoybeans,mostacreageintheU.S.andelsewhereisnowplantedwithtransgenicvarieties—butnobigcataclysmhasoccurred,reducingthetractionofthe“wehaven’tprovedit’ssafe”argument.Wecan’tproveplanesaresafe,either,yetwestillfly.

Everythinghasrisk.Whatweneedtodoisevaluatethebalancebetweenriskandbenefit.Sevenbillionpeopleneedtoeateveryday.—Interview by Nicole Kresge M

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31HHMI Bulletin / Winter 2015Jorge Dubcovsky believes transgenic regulation needs to change.

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Perspectives & Opinions

Winter 2015 / HHMI Bulletin

Dominique BergmannHHMI-GBMF InvestigatorStanford UniversityIt’simportanttothinkaboutseparatingscientificfromsocialandeconomicconcerns.Forexample,therearepeoplewhoworryaboutcorporateownershipofbiologicalmaterials;Ithinkthisisalegitimateconcern,butwehavetotalkaboutsuchthingsinaseparatebreathfromtheactualgeneticmodification.

WhenItalktopeoplewhoareanti-GMO,Itrytofigureouttheirreasoning.Ifthey’reconcernedaboutsafetyorputtingforeigngenesinfood,Iaskthemwhentheyfirsttriedquinoaoracaiberries.Usually,theysaynotuntiladulthood.Ithenaskwhytheywerewillingtotaketheincredibleriskofconsumingsomethingwithupwardsof20,000genestheyhadneverbeforebeenexposedto.Ifollowitupwithalittlelighthistoryofhowwe’vemanipulatedplantsforthousandsofyears.IdoubtthatI’veactuallyconvincedmyPaloAltoneighborstheyshouldn’tfearGMOs,butI’veatleasttaughtthemalittlescience.

Luis R. Herrera-EstrellaSenior International Research ScholarCenter for Research and Advanced Studies of the National Polytechnic Institute, Irapuato, MexicoInthe1980s,papayaproductioninHawaiiwasalmostcompletelydestroyedbythepapayaringspotvirus.Nonaturalresistancetothisdiseaseexistedinpapayas,andtheimmediatesolutionwastoapplylargeamountsofinsecticides.Usinggeneticengineering,researchersatCornellUniversitywentontodevelopgeneticallymodifiedpapayaplantsthatwereresistanttothedisease.TheseGMpapayassavedpapayaproductioninHawaii;theyalsogeneratedacleanerproductthatwasnotcontaminatedbytoxicinsecticides.ConsumersintheU.S.,Canada,Mexico,andEuropehavebeeneatingGMpapayasforthepast18yearsand,todate,notasinglehealthorenvironmentalproblemhasbeenscientificallylinkedtothisproduct,supportingthesafetyofGMcrops.

ManyotherexamplescanbeusedtoshowthatGMOsarenotonlysafe,buttheyalsohelpusdevelopsustainableagriculturewithalessnegativeenvironmentalimpact.

Michael Eisen HHMI Investigator University of California, BerkeleyThesimpleansweriseducation.OppositiontoGMOsisbornlargelyofignorance,andscientistsshouldtakeeveryopportunitytheycantoexplainthetechnologyandmotivationbehindgeneticengineeringtothepublic.

Butevenifwedothat,IthinkstrongoppositiontoGMOswillremain,becausepeopledon’tseeagoodreasonto“messwithMotherNature.”SoIthinkthemostimportantthingwecandotopromoteacceptanceofGMOsistodevelopGMOsthatoffersomethingtoconsumers,notjusttofarmers.Ifcitrusgreeningwipesouttheorange,forexample,Ithinkconsumerswillembraceengineered,disease-resistantvarieties,andresistancetothetechnologywillbegintodisappear.

Q&A How can scientists help ease society’s fear of GMOs?GMOsmakemanypeoplenervous.They’reafraidofhealthrisks,effectsontheenvironment,andwhatcouldcomeoftamperingwithnature.Butsomeofthosefearsmaybeunfounded.Here,fourscientistssuggestwaystoeaseGMOanxiety.–Edited by Nicole Kresge B

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Steven HenikoffHHMI InvestigatorFred Hutchinson Cancer Research CenterQuellingirrationalfearsoflow-probabilityevents,suchascatastrophicmeteorimpactsandadversehealtheffectsfromGMOfoods,putsscienceonthedefensive.ThedebateoverlabelingGMOproductspitsscienceagainstanti-sciencedisguisedasconsumerprotection,whentheunstatedimplicationisthatGMOsrequireahealthwarninglikecigarettes.ButthetwomajorGMOtechnologies—herbicidetoleranceandinsectresistance—havehadanundeniablypositiveimpactontheenvironment.Herbicide-tolerantcropspromoteno-tillfarmingthatreduceserosion,wateruse,andpollution.Bothtechnologiescutdownontheapplicationoftoxicchemicals,andtheaccompanyingimprovementsinyieldreducetheacreagedevotedtoagriculture.DebatingtheecologicalimpactsofGMOcropsmightmakeitmoredifficulttopromotedeceptiveeconomicandpoliticalagendas.

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34 scienceeducation ImmersionintheLab36 toolbox MicroscopesfortheMasses38 labbook Decisions,Decisions Cancer’sNicheProblem JammingJumpingGenes

Whenitcomestoprimping,fruitfliesarehardtomatch.Giventhechance,thewingedinsectswillspendupwardsof20minutesremovingdirtfromtheirbodies.Bycoveringfliesindustandwatchingthemgroom,JulieSimpson’steamattheJaneliaResearchCampusdiscoveredthattheinsectsperformameticuloussequenceofmovesthatstartsattheireyesandendsattheirthorax.Thisheatmapofafly’sheadrevealstheareasthatremaineddustyafteraminuteofgrooming,withthedustiestglowingyellow.Read more about Simpson’s research in “Habitual Grooming” on page 39.

Winter 2015 / HHMI Bulletin

Chronicle / Science Education

34 Winter2015/HHMIBulletin

Immersion in the LabAn international summer program gives talented Russian high schoolers a taste of life at the bench.biologistfyodorkondrashov’slectureonDNAataRussiansummerprogramwentbadly.He’dbeeninvitedtotalkataSoviet-styleencampmentwherethousandsofteenagersweredividedintointerestgroups:aspacesquad,asportssquad,andsoon.Kondrashovwouldbetalkingtofuturemariners.

ThesailorshadspenttheafternoonrowingontheBlackSea.“Theydockedatthepier,”recallsKondrashov,“totallytiredafterthreehoursofrowing.Theirteamleaderswerelike,

‘Andnow—alectureongenetics!’Iknewmylecturewasruined.”

Buttheyoungsailors’miseryprovedinstructiveforKondrashov:ayearlater,whenaRussianfoundationaskedhimtohelpdesignabiologysummerschoolforsuper-motivatedRussianhighschoolers,hemadesurethestudentsknewwhattheyweregettinginto.Noonewouldspringageneticslectureonthemunawares.They’dattendbecausetheywanted tospendfivehoursadayinabiologylaboratory.Furthermore,studentswouldhavesomechoice:they’drequestalabassignmentafterrotatingthroughsixofninelabs.

TheRussian-bornKondrashovhasnowservedforthreesummersasscientificdirectoratthe17-dayMolecularandTheoreticalBiologySchool,or“bioschool.”Thestudents,whopaynothingandnumberfrom60to80,comefromalloverRussia—somefromelitehighschoolsinMoscow,othersfrom“somewhereinthemiddleofSiberia.”TheygatherinAugustinPushchino,aresearchcommunityfourhoursfromMoscow.Thescientists,postdocs,andgraduatestudentswhorunthelabscomefromRussia,theUnitedStates,or,likeKondrashov,Europe;heisanHHMIinternationalearlycareerscientistattheCentreforGenomicRegulationinBarcelona,Spain.

Thestudentsworkonrealexperimentsthatadvancetheworkofthelabheads.“There’stonsofeducationliteraturewheretheytalkabouthowauthenticeducationexperienceenhanceskids’motivation,”saysKondrashov.Partofauthenticityisseeinghowscientistsrespondwhensomethinggoeswrong,hesays.“Ifyouhaveawell-rehearsedproject,nothingeverissupposedtogowrong.”

Thestudentsalsoexperience“themundanekindofeverydaythings,likedoingathousandPCRs,becauselifeinthelabcanbeveryboring.”It’skey,saysKondrashov,that“theyunderstandthattheylikedoingthemundane.”That’sespeciallyimportantinRussia,forwhenstudentsenteruniversityafter11thgrade,theirchoiceofcareerisnearlyimpossibletochange.

For15-year-oldGrigoryKhimulya,bioschoolwas“oneofthemostimportanteventsthathappenedinmylifesofar,andIfeellikethisisthecaseformostofthekids.”Now17andafreshmanatHarvard,Grigorysayshelovedworkingon“realresearchquestionswitharealprincipalinvestigator...toresearchsomethingthatnoonehadexploredbefore.”

GrigoryworkedwithJoséLuisGarcía-Pérez,anHHMIinternationalearlycareerscientistfromSpainwhostudieshumantransposableelements,or“jumpingDNA.”The10studentsonhisteamproposedtestingwhetheribuprofenwouldcauseDNAsequencesinhumanculturedcellstojumptonewpositionswithinthegenome.Theydiscoveredthatthecommonpainkillerdidseemtoincreasetherateofretrotransposition,andGarcía-Péreziscontinuingtostudytheeffect.

Alltoooften,saysGarcía-Pérez,students“don’tknowwhatscienceisuntillateincollegeoringraduateschool.”Butatbioschool,theygetafeelfor“livinginthelab.”

Theyalso“cometosee[science]asacollaborativeprocess,”saysbioschool

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bulletin/winter-2015.

“For many of them, it was the first time they’d ever talked to adults about science outside a classroom. ”—maximimakev

HHMIBulletin/Winter2015

labheadMikhailGelfand,aprominentquantitativebiologistandformerHHMIinternationalresearchscholar.“It’simportantpsychologically.”Gelfand’slablistedbioschoolstudentsasco-authorsonarecentpaper.

DoctoralstudentMaximImakaevhasnoticedastarkdifferencebetweenhowRussianandAmericanstudentsviewthechancetodoresearch.FortheBoston-areahighschoolershesupervisesattheMassachusettsInstituteofTechnology,hesayslabworkisoneamongmanyextracurricularactivities—“justanotherprogram.”Bioschoolstudents,ontheotherhand,wereecstaticaboutdoingrealresearch.“Formanyofthem,itwasthefirsttimethey’devertalkedtoadultsaboutscienceoutsideaclassroom,”saysImakev.“Itshowedstudentsthattherecanbeotheradultsintheirliveswhorespectthem,whobelieveinthem.”

Thatwasmemorablefor17-year-oldAygulMinnegalieva,who’dneverbeforedonealabexperimentbutsaidthebioschoolscientiststreatstudentslike“youareequaltothem.Theydon’tlookatyoulikeyou’rejustahighschoolstudent.”

Kondrashovsaysbioschoolvalidatesstudentswho“feeltheyareoutcastsbecausethey’resmart,becausethey’reinterestedindifferentthingsthantheirpeers.”

ThiswasarevelationforAnastasiaGoshina,17:“Iwassurroundedbypeoplewhounderstoodme.Icouldaskthemwhattheythinkonbiologicaltopics,andthey’dgivetheiropinions.We’dmaybeargueonthings,oragree.Idon’thaveopportunitieslikethatatmyschool.”Anastasia—whosegraduatingclassinTogliatti,1,000kilometersfromMoscow,hasjust20students—keepsintouchwithherbioschoollabheadandfriendsviaemailandsocialnetworking.

SheplanstoapplytouniversityinMoscow.Kondrashovsaysonemeasurableoutcomeofthesummerschoolishowmanystudentsarewillingtorelocatetoenrollinthebestprograms,sinceit’sunusualforRussianstudentstodoso.Halfthebioschoolalumninowinuniversitylefttheirhometownstofurthertheireducation.

Thebioschoolwillreconvenethissummer,fundedprimarilybytheDynastyFoundation,Russia’sfirstfamilyfoundation,andinpartbyHHMI.

But“bioschooldoesn’tendwhenthesummerends,”saysGrigory.“I’mstillpartofthenetworkofresearchersandstudents….I’minBoston,butI’mstillfinishingprojectsforlabsinMoscowandBarcelona.”– Cathy Shufro

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Chronicle / Toolbox

Microscopes for the MassesJanelia’s Advanced Imaging Center gives visiting scientists access to cutting-edge microscopes.atjanelia,scientists buildtools.Somearesmallandeasytoshare:sensorsthatcanbepurchased,softwarethatcanbedownloaded,websitesthatcanbevisited.Others,likeinnovativemicroscopes,arenot.

Thesehulkingcollectionsoflenses,mirrors,lasers,andelectronicsneedtobepreciselyalignedandboltedto800-poundopticaltables.Furthermore,manyareprototypes,meaningthatoutsidescientistslikeHernánLópez-SchierofHelmholtzZentrumMüncheninMunich,Germany,havelittleaccesstothem.

López-Schieruseszebrafishtostudynervecellsthattransmitsensoryinformationtotheorganism’sbrain.Bycuttinganaxon—athreadlikeextensionofanervecell—withalaserandwatchingitsrecoveryunderamicroscope,López-SchierlearnshowSchwanncells—whichinsulateandmaintainaxons—respondtothedamage.Withcommerciallyavailablemicroscopes,hecangetageneralideaoftheprocess.Buttoseeitindetail,heneedsamachinethatcancapturetheactionathighspeed,inalivefish,withoutdamagingitstissue.

FortunatelyforLópez-Schier,theJaneliaAdvancedImagingCenter(AIC)recentlyopeneditsdoors,offeringaccesstothemicroscopesbuiltattheAshburn,Virginia,researchcampus.“Janeliaisatool-buildingplace,andwewantourtoolstobeused,”explainsJaneliaExecutiveDirectorGerryRubin.“Ourgoalistoenablecutting-edgeworkbyprovidingpeoplewithinstrumentsthatjustdidn’texistbefore.”

TheAICisajointventurebetweenHHMIandtheGordonandBettyMooreFoundation(GBMF).Ithousesfivemicroscopes(seesidebar,“ASpectrumofTools”)andhasasupportstaffoffourscientiststohelpcarryoutexperiments.“GBMFwasalreadyinterestedinimagingandisfundingseveralgroupsdevelopingnewmicroscopes.Together,weconcludedthatprovidingwidespreadaccesstoprototypesofnewinstrumentswouldbeverycomplementarytoourongoingactivities,”saysRubin.

WheneveracallforproposalsgoesoutfromtheAIC,experimentalbiologistsanywhereintheworldcansubmitanapplicationtouseitsmicroscopes.SuccessfulapplicantsaregivenfreelodgingatJaneliaandaccesstoallthescientificfacilitiesoncampus.Thefirstroundofproposalsincludedstudyingmicrofluidicdevices,

squidhatchlings,transgenicmoss,andbacterialmembranes.

“Wewanttoencouragescientistsfromfieldsthathistoricallydon’tleverageadvancedmicroscopyintheirwork,”explainsthecenter’sdirector,Teng-LeongChew.TheAICistargetinguserswholackaccesseithertocutting-edgeimaginginstrumentsortoasupportteamversedinthelatestmicroscopytechniques.

Andbypushingthemicroscopes’experimentallimits,userscanalsoprovidevaluablefeedbacktothebuilders.“Inaway,AICbecomestheconvergentpointwherebiologymeetsengineering,”explainsChew.“It’snotjustthatJaneliaputsforwardthisreallywonderfulequipmentforsciencetouse;it’salsothatthevisitorswillinformthecontinueddevelopmentoftheinstruments.”Theirfeedbackwillhelpfurtherrefinethe

“Our goal is to enable cutting-edge work by providing people with instruments that just didn’t exist before. ”—gerryrubin


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microscopesandenabletheinstrumentstobeusedinavarietyofwaysinthelifesciences.Thehopeisthat,eventually,someAICprototypeswillbefine-tunedtoapointwheretheymightbecommercializedforbroaderdistribution.

López-SchierwasoneofthefirstscientiststousetheAIC’slatticelightsheetmicroscopetolookatafish.Butadaptinghisrelativelylargesampletothemicroscopeprovedachallenge.“Initially,wecouldn’timageanythingbecausewecouldn’tmakethefishfitwithintheholderthatwouldplaceitundertheobjectivelensesofthemicroscope,”recallsLópez-Schier.ButheandtheAICsupportteamspentacoupleofdaysmodifyingthemountingprocedureandwereabletomakeitwork.

HewasrewardedwithlivevideoofSchwanncellsrepairingdamagedaxons.Thefootagerevealedthatthecellseatupdeadaxons,aprocessknownasphagocytosis.“Withthelatticelightsheetmicroscope,wecouldactuallyseetheseintracellularvacuolesformingfromthephagocyticactivityoftheSchwanncellsastheyconsumedthedeadaxons,”heexplains.

BackinGermany,López-Schierhassubmittedapaperwithhisresults.HealsohashissightssetonbuildingasimilarmicroscopeinMunich.–Nicole Kresge


A Spectrum of Tools

Lattice light sheet microscope:InventedbyJaneliaGroupLeaderEricBetzig,thismicroscopeallowsuserstopeerinsidelivingcells,givingthemtheabilitytocreatedetailed,three-dimensionalmoviesofbiologicalprocesseslikecelldivision.(Seepage12formoreonBetzig’swork.)

Interferometric photoactivated localization microscope (iPALM): DevelopedbyJaneliaGroupLeaderHaraldHess,theiPALMisespeciallysuitedforhigh-resolutionstudiesofhowmoleculesinteractwitheachother,suchasinasignalingcomplex.

Single molecule total internal reflection fluorescence microscope (smTIRF): UsedforinvitroimagingofcomplexbiochemicalreactionslikeDNAtranscription,thesmTIRFwasdesignedandconstructedbytheresearchteamsledbyHHMIPresidentRobertTjianandphysicistStevenChuofStanfordUniversity.

Aberration-corrected multifocus microscope (acMFM): DesignedandbuiltbytheresearchteamledbythelateJaneliaGroupLeaderMatsGustafsson,theacMFMcanimageninefocalplanessimultaneously,makingitperfectfortrackingthemovementofmoleculesinsidelivingcells.

Live cell multicolor structured illumination microscope (SIM):AlsodevelopedbyMatsGustafsson,theAIC’sSIMisoneofthefastestinstrumentsofitskindandiscapableofdoingthree-dimensional,multicolor,high-resolutionimaging,makingitidealforobservingmacromolecularinteractionsandsubcellularstructuresinalivecell.

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Chronicle / Lab Book

I N B R I E F

A GROWING FAMILY TREEAbout 8,000 years ago, a band of brown-eyed, pale-skinned farmers made their way to Europe from the Near East. They mingled with the indigenous blue-eyed, swarthy hunter-gatherers, and their offspring eventually became modern Europeans. But that might not be the entire story. HHMI Investigator David Reich of Harvard Medical School has found evidence that modern Europeans have a third branch in their family tree.

Reich and a team of more than 100 collaborators worldwide compared the DNA of 2,345 modern Europeans to DNA in the remains of a 7,000-year-old farmer from Germany and eight 8,000-year-old hunter-gatherers from Luxembourg

and Sweden. “What we find is unambiguous

evidence that people in Europe today

have all three of these ancestries: early Near

Eastern farmers who

brought agriculture to Europe, the indigenous hunter-gatherers who were in Europe prior to 8,000 years ago, and ancient north Eurasians,” says Reich.

Although nearly all modern Europeans tested showed evidence of DNA from the third group—north Eurasians—the team did not find that same DNA in ancient hunter-gatherers or ancient farmers. This means the north Eurasians came to Europe after agriculture had been established—a scenario most archaeologists had believed unlikely. The scientists’ data, published September 18, 2014, in Nature, also reveals that the first Near Eastern farmers to reach Europe had ancestors from a previously unidentified lineage, which Reich’s group named the Basal Eurasians.

CRUNCHING BIG DATANowadays, microscopes capture images of the brain in unprecedented detail. But with that detail come mountains of

complex data that can slow even the fastest computer to a crawl. On a single machine, “you can load the data, start it running, and then come back the next day,” explains Janelia Group Leader Jeremy Freeman. “But if you need to tweak the analysis and run it again, then you have to wait another night.” For larger data sets, the lag time might be weeks or months.

Freeman joined with Janelia Group Leader Misha Ahrens to find another way. The scientists realized that a new distributed computing platform called Spark, which divvies up tasks across a cluster of computers, was particularly well suited to the challenges of neural data. Building on the technology, Freeman and Ahrens developed an open-source library, dubbed “Thunder,” for analyzing large-scale neuroscience data. With their library, tasks that before would take days can be completed

in hours or minutes—ideal for supporting high-throughput, exploratory analysis of large data sets.

In a report published July 27, 2014, in Nature

Methods, the Janelia team illustrated Thunder’s capabilities by using it to rapidly identify patterns of biological interest in high-resolution images of the brains of mice and zebrafish.

Thunder is designed to run on a private computer cluster or on Amazon’s cloud computing services. It is totally open source; information and tutorials can be found via the GitHub project page at freeman-lab.github.io/thunder.

HIPPOCAMPAL STORAGE SPACEThe neurons in the brain’s hippocampus store recent information about people, places, and events. But how does the hippocampus avoid running out of storage space P

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Decisions, DecisionsA stress hormone can flip the switch between strategic and random behavior in rats.we’rehardwiredto learnfromourmistakes.Butsometimesthebrainabandonslessonslearnedandactslessstrategically.Inthewild,forexample,animalsoftenmoveunpredictablytoescapepredators.Howthebrainswitchesbetweenpayingattentiontopastexperiencesandproceedingrandomlyisclearernow,thankstonewfindingsfromJaneliaGroupLeaderAllaKarpova.

“Normally,wedeterminethebestcourseofactionstrategically,bymakingamodelofthe

worldbasedonalltheinformationavailabletous—inotherwords,ourexperience,”explainsKarpova.“Butsometimesitcanbebetternottouseall—or,forthatmatter,any—ofthatinformation.”

KarpovaandpostdocGowanTervocreatedagameinwhichratswerepresentedwithtwoholes,onlyoneofwhichcontainedfood.Thefood’slocationwasdeterminedbyoneofthreeincreasinglytoughcomputerprogramsthatlearnedfromtheratandattemptedtothwartitsstrategy.Thefirstwaspredictableandfairlyeasytobeat,thesecondabittrickier,andthethirdessentiallyunbeatablebecausetheprogramusedaverysophisticatedpredictionalgorithmthatwashardfortheanimaltocounter-model.

Theanimalsusedastrategicapproachwhenplayingtheeasyopponent.Butwhentheywentupagainstthesophisticatedcompetitor,theystoppedbasingchoicesonpreviousexperience,andtheirdecisionsbecamehaphazard.Infact,manyoftheratsgotstuckinrandommodeforhundredsoftrials,evenwhenthefoodbecameeasytofind—aneffectresemblingapsychologicalconditioncalledlearnedhelplessness.

AsKarpovaandTervoreportedSeptember25,2014,in Cell,theydiscoveredthatalteringlevelsofastresshormonecallednorepinephrineinanareaofthebraincalled

theanteriorcingulatecortexchangedtherats’operatingmode.Increasingnorepinephrinesuppressedtherats’strategicmodeandcausedthemtobehaverandomly;inhibitingreleaseofthehormonehadtheoppositeeffect.

“Justbymanipulatingasingleneuromodulatoryinputintoonebrainarea,youcandramaticallyenhancethestrategicmode,”saysKarpova.“Theeffectisstrongenoughtorescueanimalsoutoftherandommodeandsuccessfullytransformthemintostrategicdecisionmakers.”

Karpovasuspectsthatthesameneuralmechanismsgovernthewayhumansact,whichcouldleadtoafuturetherapyforlearnedhelplessness.– Nicole Kresge

Random behavior often helps animals escape predators.

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if, say, an environment is larger than predicted or an experience goes on longer than expected? According to Janelia Group Leader Albert Lee, it’s all about division of labor.

Lee and his team created a 48-meter-long maze and recorded which neurons in the hippocampus fired when a rat explored a small area of the maze for the first time. Then the researchers incrementally increased the portion of the maze the animals could access and monitored how the brain added the new information to its spatial map.

As they reported August 15, 2014, in Science, the team discovered that each neuron contributes to the map at its own rate. As the rat explored the maze, some cells fired early and often and associated themselves with the new space immediately, while others held back. These slower cells rarely contributed unless the space expanded beyond the original size captured by the early-firing neurons.

“Instead of the hippocampus having to adjust in time as the animal

notices that the maze gets larger, it anticipates all different sizes of mazes from the beginning,” Lee says. “It does this by dividing up its population of neurons so that certain cells are ready to represent smaller mazes, others are ready to represent medium-size mazes, and still others, large mazes.” Lee believes that similar mechanisms are at work when the human brain records a new experience.

HABITUAL GROOMINGFruit flies are very fastidious when it comes to cleanliness. A dust-covered fly may spend upwards of 20 minutes removing grit and grime in a habitual sequence that starts at its eyes and ends at its thorax. Studying this structured grooming has helped Janelia Group Leader Julie Simpson explain how the brain organizes sequential behavior.

“You can’t do everything at once,” says Simpson. “How do you manage competing demands on your brain and

limbs so that you execute things in order of importance?” As she reveals in the August 19, 2014, issue of eLife, it’s all about hierarchy.

Simpson, postdoctoral researcher Andrew Seeds, and their colleagues devised a way to trigger particular grooming tasks by turning on different sets of neurons. They discovered that dust-covered flies always started their grooming program at the beginning—cleaning their eyes—but weren’t able to progress beyond the point that the scientists switched on. For example, when the researchers activated neurons for abdominal cleaning, the fly would clean its head and then its abdomen but then would continue cleaning its abdomen, ignoring the dust on the rest of its body.

“It progresses through the hierarchy to that particular

point, and then it gets stuck,” Simpson

explains. This means that earlier grooming

steps have the ability to

halt those that happen later. These findings may help explain other behaviors that involve sequences of tasks, like nest building in birds.

SENSITIVE SEQUENCINGTechniques known as Sanger sequencing and whole exome sequencing are great tools for identifying defective genes that are present in all of an individual’s cells. But they aren’t as reliable when mutations occur in just a few cells. Although not as common, these “mosaic” mutations are increasingly recognized as a cause of disease, especially in the brain. “If a mutation is in only 5 or 10 percent of the cells, then that’s going to be in a very small fraction of the data, which will be hard to separate from the noise,” explains HHMI Investigator Christopher Walsh of Boston Children’s Hospital.

To overcome that challenge, Walsh and his colleagues developed a new strategy that flushes out these hard-to-find mutations. The team focused on a set of genes associated with brain P

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Cancer’s Niche ProblemIn some bone marrow cancers, a stem cell’s surroundings play a surprising role.untilnow,scientistsbelievedthatcancerwasdrivenbychangesinsidecells:genesbecomemutated,cellsgrowunchecked,andatumorforms.Buttheremaybemoretothestory.NewresearchfromHHMIInternationalEarlyCareerScientistSimónMéndez-Ferrerindicatesthat,insomecases,whatgoesonjustoutsideacellalsoplaysaroleincancerformation.

Inagroupofbonemarrowdisordersknownasmyeloproliferativeneoplasms,adefective

genecauseshematopoieticstemcells(HSCs)tomaketoomanybloodcells.Asthebloodcellsbuildup,thediseaseworsens,sometimesevolvingintocancer.Theonlyrealcureisabonemarrowtransplant.However,sometransplantpatientsdevelopanewbonemarrowcanceroriginatingfromthedonortransplantcells,hintingthatsomethingotherthanthepatient’smutantHSCsaretoblame.

Studyingtheprocessinmice,Méndez-Ferrer’steamdiscoveredthatthemutantHSCsproducedanabundanceofsmallinflammatoryproteinscalledcytokinesthatweredamagingnearbyneurons.Thisdamage,inturn,preventedthenervesfromactivatingothercellsthathelpregulateHSCs.Theendresult:uncheckedgrowthofHSCsandthepotentialofmyeloproliferativeneoplasms.

“Eventhoughthediseaseisinitiatedbyamutatedbloodstemcell,thisstemcellcannotdrivethediseaseuntilitdestroysparticularelementsofthemicroenvironment,”explainsMéndez-FerrerattheSpanishNationalCenterforCardiovascularResearch.“First,ithastodamagethenervecellsinthemarrow.”

Whentheresearchersaddedananalogoftheneurotransmitteradrenalinetocompensateforthedamagedneurons’inabilitytofire,themousecellswereonceagainabletocontrolHSCproliferation.TheteamreporteditsfindingsAugust7,2014,inNature.

Mosteffortsattreatingmyeloproliferativeneoplasmsarefocusedonblockingthe

mutantHSCs,accordingtoMéndez-Ferrer.“Weshowedthatanalternativeapproachmightbetoprotecttheniche—theenvironment—andcontroltheexpansionofthemutatedcell,”hesays.Withthatinmind,Méndez-Ferrer,RadekSkoda,andothercolleagueswillsoonstartaclinicaltrialtoseeifadrenalineanaloguescanbeusedtoeffectivelytreatpatientswithmyeloproliferativedisease.– Nicole Kresge

The environment around these hematopoetic stem cells plays a role in cancer formation.

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malformations. They isolated DNA from patients with unexplained brain malformations and then sequenced each of these genes hundreds to thousands of times.

“We said we’d shoot to sequence them a thousand times over,” Walsh says, “thinking that if a mutation is only present in 5 percent of the cells, it will be obvious that it’s a mutation, because we’ll see that mutation 50 times.” The team reported August 21, 2014, in the New England Journal of Medicine that they had found disease-causing mutations in more than a quarter of the patients in their study, demonstrating the effectiveness of their method and further supporting the idea that mosaic mutations are an important cause of diseases that affect just a part of the body.

APPETITE CONTROLSometimes science is serendipitous. Researchers—like HHMI Investigator David Anderson—occasionally find something they weren’t looking for. In Anderson’s case, the accidental

discovery was a few thousand cells that turn appetite on and off.

Harnessing a technique called optogenetics, Anderson and his colleagues at the California Institute of Technology used a beam of light to activate a small group of neurons in a part of the mouse brain called the amygdala. In earlier research, Anderson had pegged these nerve cells as part of the fear response. But, as the team reported September 2014 in Nature Neuroscience, rather than eliciting fearful or anxious behavior, the light caused the mice to abruptly stop eating. The scientists were able to rule out the possibility that the animals were too scared to eat, and they also established that the neurons were also activated by bad tastes, nausea, and feelings of satiety.

“We think the neurons make up a central node that integrates the influences of multiple [feeding-inhibitory] signals and relays this information to inhibit other brain centers that normally promote feeding,” explains Anderson. Many

patients with eating disorders also have emotional disorders like depression and anxiety, which may be explained by the neurons’ dual appetite and fear functions. Next, Anderson wants to examine how the nerve cells manage to regulate both feeding and emotional behaviors.

QUALITY PATROLLike intricate pieces of origami, proteins need to be folded in just the right way. An incorrect bend here or an extra loop there can render a molecule useless—or even toxic—to the body. To dispose of these defective proteins, cells detect misfolded proteins and tag them with a protein called ubiquitin, marking them for degradation.

In the yeast Saccharomyces cerevisiae, there are two well-established roving protein complexes that add ubiquitin to proteins in the endoplasmic reticulum (ER). One complex is for proteins whose misfolded domains occur in the cytosol, and one is for

proteins with misfolded domains in the lumen or membrane of the

endoplasmic reticulum. International Early Career

Scientist Pedro Carvalho recently uncovered a third complex that targets proteins associated with a specialized ER domain in the inner nuclear membrane (INM).

When Carvalho and his team at the Center for Genomic Regulation in Barcelona, Spain, removed the two known protein patrol complexes from yeast, they noticed that some molecules were still being disposed of, suggesting that another scout was at work. As they reported September 18, 2014, in Science, this mystery warden turned out to be the Asi complex—a gang of three molecules in the INM that not only gets rid of misfolded proteins, but also removes certain correctly folded proteins that are incorrectly directed to the INM.

Carvalho and his team are now looking for other proteins that bind to this patrol complex.


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Jamming Jumping GenesCells have a rapidly evolving arsenal to counter wayward DNA.formillionsofyears ,bitsofDNAcalledretrotransposonshavebeenduplicatingandinsertingthemselvesrandomlythroughoutthegenomesofdifferentorganisms,includinghumans.Sometimesthejumpinggenesalightinplacesthathelpspeciesevolvenewtraits.Othertimestheydonothing.Butalltoooftentheytouchdowninthemiddleofagene,alteringthewayit’sregulatedorknockingitoutofcommission.Fortunately,asHHMIInvestigatorDavidHausslerrecentlyshowed,cellshaveevolvedawaytokeepthesewaywardsequencesincheck.

“Overgenerations,ourgenomebecomesbloatedwithcopiesofretrotransposons,”explainsHaussler.“Theydodamagebydisruptingnormalgeneticmechanisms.

Somethinghastobethesecuritypatroltotryandshutthesethingsdown.”

Inhumans,oneformofsecuritypatrolcomesfrommorethan400rapidlyevolvinggenesthatproducewatchdogproteinscalledKRABzinc-fingers.Thesewatchdogsscanthegenome,clampontoretrotransposons,andthencallonotherproteinstosilencethem.

ToseeKRABzinc-fingersinaction,Haussler’steamattheUniversityofCalifornia,SantaCruz,usedmousecelllinescontainingacopyofhumanchromosome11,whichincludeshundredsofretrotransposonscommonlyfoundinprimateslikehumans,butnotpresentinrodents.Sincemicehaverodent-specificKRABzinc-fingerstocontroltheirretrotransposons,therodentcellscouldn’tstoptheroguehumanretrotransposonsfromexpressingthemselves.Thatis,untiltheresearchersaddedtwohumanKRABzinc-fingers—ZNF91andZNF93—thathaltedtheprimate-specificretrotransposonsintheirtracks.TheteampublisheditsfindingsSeptember28,2014,onlineinNature.

HausslerexplainsthatmutationsinretrotransposonsallowthemtoescapedetectionbytheKRABzinc-fingers,whichinturndrivestheevolutionofnewKRABzinc-fingergenes.“Byreconstructingmolecularevolutionaryhistories,wecanseethatthese

KRABzinc-fingergeneshavebeenmajorplayersinthisbattlewiththeretrotransposonsandwillprobablycontinuetobeso,”hesays.

ThereisasilverlininginthisseeminglyendlessarmsracewithinourownDNA.OnceKRABzinc-fingersarenolongerneededtosuppressretrotransposons,manyofthesewatchdogproteinsadoptnewrolesasregulatoryproteins,controllingtheactivityzofgenesnearretrotransposonlandingsites.– Nicole Kresge

Primates have evolved specialized weaponry to home in on bits of jumping DNA.

12Immune cells will do anything within their power to do their jobs. Some, like this neutrophil, are driven to crawl, swim, or ooze through our bodies’ tissues and fluids to be a first responder at an acute infection or inflammation. Until recently, we couldn’t have known what these cells look like in action, as they morph from a sphere into a shape-shifting tangle of appendages and ruffled edges. But now, remarkable advances in imaging live cells offer a whole new view of biology.

HHMI Bulletin / Winter 2015

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The Essence of our BeingWe humans may think of ourselves as solid objects, all flesh and bone. But take a close look, and it’s clear our bodies are composed largely of oxygen and hydrogen. We are essentially ephemeral—akin as much to wind, water, and fire as to earth. No matter how heavy you feel, says biologist and author Curt Stager, you are more like a “porous froth of atomic Styrofoam” than a dense mass. Understanding the world in atomic terms can give us a richer appreciation, he says, for how we interact with each other and the environment around us.

What do atoms have to do with you? Everything. They were present and intimately involved when you and everyone you have ever loved—or hated—did everything that you and they have ever done. Every scent you’ve ever savored, every sight you’ve ever seen, every song you’ve ever enjoyed, every cry or sigh that ever passed your lips sprang from atoms at work within the atmosphere and the darkest recesses of your body. When you eat, the bodies of other living things become part of you. If you cut yourself, the wreckage of dying stars runs out in a stream of ancient atoms that triggered some of the most violent explosions in the cosmos. When you flush your wastes, you scatter the atomic echoes of lightning bolts and volcanoes into a global cycle that may some day return them to you, as unpleasant as that may sound. And whenever you grin, the sparkling of your teeth conceals the dim afterglow of nuclear fallout from Cold War bomb tests over the Pacific.

You are not only made of atoms; you are atoms, and this book, in essence, is an atomic

field guide to yourself. All you need in order to interpret your life in primal elemental terms is access to some of the latest scientific information, some new ways to reconsider your world in light of it, and an active imagination. In doing so you will begin to experience a revolution in self-awareness that is playing out on a larger scale around the world.

From Your Atomic Self by Curt Stager. Copyright ©

2014 by the author and reprinted by permission of

Thomas Dunne Books, an imprint of St. Martin’s

Press, LLC.Lil

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bulletinWinter ’15 Vol. 28 No. 1

in this issue Allergy, Asthma, and the Gut

Germline Stem CellsScience Posses Fan Out

Super Resolution

Nobel-winning work provides a stellar

view of cellular detail

Cradle of LifeMystery and awe surround the genesis of life, whether observed

from afar or through the lens of a microscope. Even in the ovary of a fruit fly, as seen here, the beauty and wonder are undeniable.

The orb-like organ is where primordial germ cells, or PGCs (green), rub shoulders with special “intermingled cells” (red), contact that is

required for the germ cells to proliferate. Later in development, some of the PGCs will become germline stem cells—destined to start the process all over again—while the others will differentiate into eggs.

Learn more about how germline stem cells hold the key to survival of a species, including humans, in “Perfect Balance,” page 18.

4000 Jones Bridge Road Chevy Chase, Maryland 20815-6789www.hhmi.org

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