Gene ExpressionGene ExpressionWhat determines the mix of What determines the mix of

proteins in a cell at any given proteins in a cell at any given time?time?

FearsHow long you

will live

How you will respond to stress

Can some Can some acquired acquired traits be traits be

passed on?passed on?

If so how?If so how?

How could acquired traits be passed How could acquired traits be passed on and can they be changed during on and can they be changed during

your life?your life? Basically, you not only pass on your genes Basically, you not only pass on your genes

to the next generation, you pass on some to the next generation, you pass on some of the signals attached to the DNA that of the signals attached to the DNA that control the reading of the genes, which control the reading of the genes, which can change the expression of a traitcan change the expression of a trait

Some of these signals can be changed Some of these signals can be changed throughout lifethroughout life

Some of these signals are reset in the Some of these signals are reset in the sperm and eggsperm and egg

Epigenetics Epigenetics Switching genes on and offSwitching genes on and off

Why do cells need to control gene Why do cells need to control gene expression?expression?

Different stages of development

Differentiation

Meet demands of the organ

Respond to changing environment

Respond to outside signals Make sure you are not wasting energy

making things you don’t need

Eukaryotic Gene ExpressionEukaryotic Gene Expression 6 x 109 base pairs ~ 35,000 genes

~5-20% of genes expressed/cell 1000-7000 genes transcribed/cell

Promoter marks the beginning of each gene, but how does cell know what genes to transcribe?

What else controls how much protein and which proteins are created in a cell and what the final protein looks like?

You are what you eat?!You are what you eat?!Or even what your mother ateOr even what your mother ate

Why Genes Aren’t DestinyWhy Genes Aren’t DestinyExceptsExcepts

“…“…conditions in the womb could affect your health not conditions in the womb could affect your health not only when you were a fetus but well into adulthood.”only when you were a fetus but well into adulthood.”

“…“…if a pregnant woman ate poorly, her child would be at if a pregnant woman ate poorly, her child would be at a significantly higher than average risk for cardiovascular a significantly higher than average risk for cardiovascular disease as an adult.”disease as an adult.”

“…“…kids who went from normal eating to gluttony in a kids who went from normal eating to gluttony in a single season produced sons and grandsons who lived single season produced sons and grandsons who lived shorter lives…. A single winter of overeating as a shorter lives…. A single winter of overeating as a youngster could initiate a biological chain of events that youngster could initiate a biological chain of events that would lead one’s grandchildren to die decades earlier would lead one’s grandchildren to die decades earlier than their peers did.”than their peers did.”

Excepts ContinuedExcepts Continued

“…“…exposed mice with genetic memory problems to an exposed mice with genetic memory problems to an environment rich with toys, exercise, and extra attention. environment rich with toys, exercise, and extra attention. These mice showed significant improvement in long term These mice showed significant improvement in long term potentiation (key to memory formation). Surprisingly, potentiation (key to memory formation). Surprisingly, their offspring also showed LTP improvement, even their offspring also showed LTP improvement, even when the offspring got no extra attention.”when the offspring got no extra attention.”

“…“…sons of men who smoke in prepuberty will be at sons of men who smoke in prepuberty will be at higher risk for obesity and other health problems…”higher risk for obesity and other health problems…”

““In 2008, the NIH would pour $190 million into how and In 2008, the NIH would pour $190 million into how and when epigentic processes control genes.”when epigentic processes control genes.”

Gene Expression Can be Controlled at Any Gene Expression Can be Controlled at Any Level of Protein Production or ActivationLevel of Protein Production or Activation

Control of Eukaryotic Gene Control of Eukaryotic Gene ExpressionExpression

I.I. Availability of genes to RNA Availability of genes to RNA Polymerase and transcriptionPolymerase and transcription factorsfactors

1. If DNA is too compact – no gene expression1. If DNA is too compact – no gene expression• Euchromatin – normal chromatin – gene Euchromatin – normal chromatin – gene

expressionexpression• Heterochromatin – more condensed – no Heterochromatin – more condensed – no

expressionexpression• Chromosomes – extremely condensed – no Chromosomes – extremely condensed – no

expressionexpression

Difference in Euchromatin and Difference in Euchromatin and HeterochromatinHeterochromatin

Dark staining areas are heterochromatin, light are euchromatin – more euchromatin – more active the cell is – always heterochromatin around the inside of the nuclear membrane

Inactive lymphocyte on leftInactive lymphocyte on leftActivated lymphocyte on the rightActivated lymphocyte on the right

Availability of DNA to RNA Availability of DNA to RNA Polymerase ContinuedPolymerase Continued

2.2. How close DNA is to the nucleosome and How close DNA is to the nucleosome and if if it is attached to the nuclear matrix it is attached to the nuclear matrix determines access of enzymesdetermines access of enzymes

3.3. AcetylationAcetylation of histone proteins unwinds of histone proteins unwinds the the DNA so enzymes have accessDNA so enzymes have access

4.4. MethylationMethylation – inhibits DNA expression, – inhibits DNA expression, responsible for DNA imprinting (methylation responsible for DNA imprinting (methylation patterns are often messed up in cancer cells)patterns are often messed up in cancer cells)

http://learn.genetics.utah.edu/content/epigenetics/rats/

Availability of DNA to RNA Availability of DNA to RNA Polymerase ContinuedPolymerase Continued

II.II. Transcriptional Transcriptional ControlControl

1.1.Controlled by Controlled by regulatory regulatory proteins and proteins and transcription transcription factors (Make up factors (Make up transcription transcription initiation initiation complex)complex)

Eukaryotic Genes – Regulatory Eukaryotic Genes – Regulatory ElementsElements

/silencer

Activators or Repressors – Bind to distal control elements

Transcription Factors – Binds to proximal control elements AND to the promoter.

DNA – Proximal Control ElementsDistal Control Elements – Enhancers and

Silencers

Proteins – Transcription Factors (general and specific)ActivatorsRepressors

Movie of Transcription Initation Complex

Genes of same enzymatic pathway are Genes of same enzymatic pathway are spread out all over the genome – so spread out all over the genome – so

how are they all expressed at the same how are they all expressed at the same time?time?

Each gene has its own promoter but many may have same proximal and distal control elements so 1 type of transcription factor and activator may control many

genes (in the same enzymatic pathways)

Another Way Transcription Is Another Way Transcription Is Controlled Controlled

2.2. Steroid HormonesSteroid Hormones

Steroids bind to a receptor and translocate Steroids bind to a receptor and translocate into nucleus. into nucleus.

Steroid/receptor complex binds to DNA in Steroid/receptor complex binds to DNA in upstream regulatory elements to switch on upstream regulatory elements to switch on genes.genes.

III.III. Control of RNA ProcessingControl of RNA Processing• Alternative Splicing – Don’t know what Alternative Splicing – Don’t know what

controls the splicing – spliceosomes bind to controls the splicing – spliceosomes bind to ends of introns but what identifies what are ends of introns but what identifies what are the introns???the introns???

• Some genes have many alternate forms due Some genes have many alternate forms due to splicingto splicing

IV.IV. Control of RNA DegredationControl of RNA Degredation1.1. Untranslated trailer of mRNA controls if Untranslated trailer of mRNA controls if

translation lasts hrs. or weeks (may be a translation lasts hrs. or weeks (may be a binding site for ncRNA’s)binding site for ncRNA’s)

2.2. ncRNA (non-coding RNA’s)ncRNA (non-coding RNA’s)

a. Micro RNA’sa. Micro RNA’s

Transcribed, folds, piece is cut off by a Dicer Transcribed, folds, piece is cut off by a Dicer which which also destroys the 2also destroys the 2ndnd strand. Single strand strand. Single strand combines with protein and then binds to and combines with protein and then binds to and inactivates mRNA’s with complementary sequences inactivates mRNA’s with complementary sequences or or causes their destructioncauses their destruction

b. Small Interfering RNA (same as miRNA but from b. Small Interfering RNA (same as miRNA but from longer pieces of RNA)longer pieces of RNA)

Both mi and si RNA can recruit enzymes to form Both mi and si RNA can recruit enzymes to form heterochromatin and therefore turn off genesheterochromatin and therefore turn off genes

How miRNA worksHow miRNA works

RNAi Video

V.V. Control of TranslationControl of Translation

1.1. Regulatory proteins can prevent mRNA Regulatory proteins can prevent mRNA from binding to the ribosome – must be from binding to the ribosome – must be removed before translation occursremoved before translation occurs

2.2. Initiation Factor (helps mRNA bind to Initiation Factor (helps mRNA bind to ribosome) ribosome) Example – fertilization turns on initiation factor so

get quick translation – prevents translation until activate the initiation factor

VI.VI. Post-Translational ModificationsPost-Translational Modifications

• Can prevent correct modifications or transportCan prevent correct modifications or transport• Can be modified differently by different Can be modified differently by different

enzymes in the rough ER of different kinds of enzymes in the rough ER of different kinds of cells (maybe different enzymes in different cells (maybe different enzymes in different RER of different cell types?)RER of different cell types?)

VII.VII. Protein DegredationProtein Degredation (Proteosome chops up unneeded proteins – (Proteosome chops up unneeded proteins – tagged with ubiquitin) Don’t know what tagged with ubiquitin) Don’t know what controls speed of degredationcontrols speed of degredation

VIII.VIII. Gene sequences Gene sequences themselves may affect expression themselves may affect expression 1.1. Multi-Gene Families - may have multiple copies Multi-Gene Families - may have multiple copies

of the same gene (can be expressed at once or of the same gene (can be expressed at once or at different times in response to the at different times in response to the environment)environment) Examples Examples

• genes to make rRNA (100-1000 repeats of these genes genes to make rRNA (100-1000 repeats of these genes together)together)

• Genes to make hemoglobin proteins – multiple copies are Genes to make hemoglobin proteins – multiple copies are slightly different – produced at different times during slightly different – produced at different times during development.development.

2. Transposons 2. Transposons “Jumping Genes” – May copy “Jumping Genes” – May copy and move or just move. May jump into a gene and move or just move. May jump into a gene and disrupt it; may jump into a regulatory area and disrupt it; may jump into a regulatory area and increase or decrease production of that and increase or decrease production of that proteinprotein

InsertionInsertion – codes for an enzyme that cuts it out – codes for an enzyme that cuts it out

ComplexComplex – has other genes that move – has other genes that move tootoo

Retrotransposons Retrotransposons – code for RNA which is – code for RNA which is then then copied into DNA and inserted somewhere else copied into DNA and inserted somewhere else

in genomein genome

Gene Sequences Controlling Expression ContinuedGene Sequences Controlling Expression Continued

3. Satellite DNA (short sequences repeated 3. Satellite DNA (short sequences repeated many times – short tandem repeats) many times – short tandem repeats)

Makes up 10-15% of genome (telomeres Makes up 10-15% of genome (telomeres are satellite DNA)are satellite DNA)

• Can act as transposonsCan act as transposons• Can be extended causing genes to malfunctionCan be extended causing genes to malfunction

4. Interspersed Repetitive DNA (25-4. Interspersed Repetitive DNA (25-40% of 40% of genome) 100-1000 b.p genome) 100-1000 b.p spread spread throughout – most are throughout – most are transposonstransposons

NeurofibromatosisNeurofibromatosis

Cause of NeurofibromatosisCause of NeurofibromatosisNeurofibromatosis 1 (1/3000) births

Mutation in the neurofibromin gene on chromosome #17

Neurofibromin is a tumor supressor gene which inhibits the p21 ras oncoprotein

It causes neural tumors all over the body due to uncontrolled cell division

Have also found cases where the disease is caused by an Alu sequence – jumping gene – jumps into the gene and messes it up

Alu sequence inserted in an intron of the NF1 gene

The presence of the Alu sequence caused a splicing error, which in turn caused one of the exons to be left out of the transcribed mRNA, thereby leading to a shift in the reading frame and production of an abnormal protein.

Researchers have reported a growing number of Alu-disease associations in disorders ranging from hemophilia to breast cancer. According to one estimate, about 0.4% of all human genetic disorders are caused by or associated with Alu

11% of genome is Alu sequences

5. Gene Rearrangements5. Gene RearrangementsExample: antibodiesExample: antibodies

6. Gene Amplification6. Gene Amplification – making more – making more copies of a gene when neededcopies of a gene when neededExample – rRNA genes in developing embryo Example – rRNA genes in developing embryo of amphibians – extra copies are small of amphibians – extra copies are small circular pieces that are destroyed after circular pieces that are destroyed after embryonic developmentembryonic developmentExample – Cancer cells in response to drugs Example – Cancer cells in response to drugs (amplify drug resistance genes)(amplify drug resistance genes)

7. Selective Gene Loss7. Selective Gene Loss – in developing – in developing embryo – only in some cells – in insectsembryo – only in some cells – in insects

Gene Expression and Gene Expression and Embryonic DevelopmentEmbryonic Development

Cytoplasmic Determinants – RNA and Cytoplasmic Determinants – RNA and proteins unevenly distributed in the egg that proteins unevenly distributed in the egg that signal developmentsignal development

Induction – signals from surrounding cells Induction – signals from surrounding cells that control developmentthat control development

Master regulatory genesMaster regulatory genes

turn on tissue specific genesturn on tissue specific genes

to make tissue specific proteins to make tissue specific proteins

Body Plan PatterningBody Plan Patterning Both cytoplasmic determinants and Both cytoplasmic determinants and

inductive signals help set up positional inductive signals help set up positional informationinformation

Homeotic Genes Homeotic Genes contain program for development of the body contain program for development of the body

plan plan Are highly conserved and have same Are highly conserved and have same

sequences within the genes called sequences within the genes called homeoboxes. homeoboxes.

Code for transcription factors and are master Code for transcription factors and are master control genescontrol genes

Homeobox GenesHomeobox Genes

Homeobox – 180 nucleotide segment of homeotic genes. Conserved in all animals – part of gene that codes for part of protein that binds to the DNA.

Hox genes found in clusters on the chromosomes. Genes are lined up in order of what part of the body they control the formation of.

Mutated Hox GenesMutated Hox Genes

OncogenesOncogenes Oncogenes – cancer causing genes Oncogenes – cancer causing genes

induced by virusesinduced by viruses

Protooncogenes – normal genes that Protooncogenes – normal genes that become cancer causing when they are:become cancer causing when they are:

• MutatedMutated• AmplifiedAmplified• Or move into an area with an active promoterOr move into an area with an active promoter• Examples – growth factors, cell cycle proteins, Examples – growth factors, cell cycle proteins,

tumor supressors (inactivated and usually supress tumor supressors (inactivated and usually supress growth), cell attachment proteinsgrowth), cell attachment proteins

Proto-oncogene ExampleProto-oncogene Example

Ras gene – codes for a G-Protein Ras gene – codes for a G-Protein (activated by a receptor when messenger (activated by a receptor when messenger binds and sets off a series of chemical binds and sets off a series of chemical reactions leading to increase in cell reactions leading to increase in cell division thru the activation of transcription division thru the activation of transcription factors)factors)

Mutated Ras gene – G protein is always Mutated Ras gene – G protein is always on even when nothing is bound to receptoron even when nothing is bound to receptor

Found in 20-25% of all human tumorsFound in 20-25% of all human tumors

Tumor Supressor GenesTumor Supressor Genes Repair damaged DNARepair damaged DNA Control cell adhesionControl cell adhesion Inhibit the cell cycleInhibit the cell cycle Activate cell suicide if damage is unfixableActivate cell suicide if damage is unfixableExample – p53 gene (p53 mutation found Example – p53 gene (p53 mutation found

in over 50% of human tumors)in over 50% of human tumors)In response to damage it :In response to damage it :

Halts the cell cycleHalts the cell cycle Turns on DNA repair enzymesTurns on DNA repair enzymes Activated apoptosis if damage cannot be Activated apoptosis if damage cannot be

repairedrepaired

Cancer Cells vs. NormalCancer Cells vs. Normal

BACTERIAL GENE EXPRESSIONBACTERIAL GENE EXPRESSION

Allows bacteria to live in a changing Allows bacteria to live in a changing environment.environment.

Operons – a whole gene unit – all genes Operons – a whole gene unit – all genes necessary for an enzymatic pathway are necessary for an enzymatic pathway are lined up behind a promoter and operator.lined up behind a promoter and operator.

Promoter Operator Genes Term. Seq.Promoter Operator Genes Term. Seq.

Bacterial OperonsBacterial Operons

Operator – controls access to promoter for RNA polymerase

Operator is always “on” unless a protein is bound to it

Repressor – binds to operator and blocks RNA polymerase from binding – specific to the operon.

Repressors for OperatorsRepressors for Operators

For Anabolic OperonsFor Anabolic Operons

Repressible OperonRepressible Operon

Product shuts off operon by activating Product shuts off operon by activating the repressorthe repressor

For Catabolic OperonsFor Catabolic Operons

Inducible OperonInducible Operon

Substrate turns on operon by Substrate turns on operon by deactivating the repressordeactivating the repressor

Repressor ConceptRepressor Concept

Tryptophan Operon: An example of an Tryptophan Operon: An example of an operon that codes for enzymes in an operon that codes for enzymes in an

anabolic pathwayanabolic pathway

The lac operon – an example of The lac operon – an example of an operon coding for enzymes an operon coding for enzymes

in a catabolic pathwayin a catabolic pathway

3-D View of the Lac Repressor3-D View of the Lac Repressor

Additional ways to activate bacterial Additional ways to activate bacterial genesgenes

Proteins bind to the promoter making it easier for Proteins bind to the promoter making it easier for RNA Polymerase to bindRNA Polymerase to bind

Example: only want to make enyzmes to break Example: only want to make enyzmes to break down lactose if lactose is present and if there is down lactose if lactose is present and if there is low glucose availablelow glucose available

Low glucose – high AMPLow glucose – high AMP AMP + CAP (catabolic activator) together bind to AMP + CAP (catabolic activator) together bind to

the promoter helping polymerase to attachthe promoter helping polymerase to attach CAP regulates several metabolic pathwaysCAP regulates several metabolic pathways