Lipid signaling

• Lipid structures– Phospholipids– Sphingolipids

• Lipid modifying enzymes• Second messenger effectors

– DAG, PA– IP3– Arachidonic acid

Phospholipids

• Glycerol Backbone• 2 hydrophobic acyl tails• Large polar “head group”• Cholesterol

Hydrophobic core

Solvent cloud

Polar transition

Sphingolipids

• Amine backbone with integral acyl• Polar headgroup• One extra fatty acid tail, usually saturated

Lipid modifiers

• Phospholipids– PLA– PLC– PLD

• Sphingolipids– Ceramidase– Sphingomyelinase– No PLD equivalent

Phospholipase C

• PI-directed lipases– Dependent on membrane association– Frequently Gq; sometimes phosphorylation

• PLC • PIP2 DAG + IP3

– Ca2+– PKC

• Reduces PIP2

– Block synthesis of PIP3

– Block PI3-K

PLC

• Membrane localization– G binding (50 nM)– Pleckstrin homology (PI-3P binding)

• Frequently nuclear, stimulating DNA synth– This mode is ERK 1/2, not Gq, dependent

• Deactivation– PLC:Gq ATPase activity ~50x native Gq– RGS

PLC C-terminal dimer binds 2 GqSinger et al., 2001

PLC

• Membrane localization– SH2 domain– PI(3,4,5)P3

• Activated by RTK mediated phosphorylation– PI-3K facilitated recruitment antagonizes PI-3K

PLC, PLC

• Poorly characterized• Delta

– Highly Ca2+ dependent– PI (3,4,5) P3/PI (4,5) P2

– Gh (thromboxane, oxytocin)• Epsilon

– Ras binding– No PH domain

IP3

• Endoplasmic reticulum IP3 channel– IP3 gated– Ca2+ activated

Protein Kinase C

• S/T kinases regulated by membrane association (and sometimes Ca2+)

• Conventional ()– Ca2+– DAG/PMA

• Novel ()– DAG– Unsaturated fatty acid

• Atypical ()– PS/PI/unsaturated fatty acid

cPKC

• Membrane targeted by DAG– Phosphorylation dependent– Ca2+ enhanced, PS

• Phosphorylation– Thr497/500– Autophosphorylation– RTK

• Inhibition– Sphingosine

Liu &Heckman 1998

Localization

• Isoform specific subcellular localization– Sensory axon terminals– Post-synaptic dendrites– Nuclear membrane

• RACK– Receptor for activated C-Kinase– AKAP

Effects

• Response– Growth– Apoptosis– Motility

• Substrates– Histone 3– Myelin basic protein– Nuclear lamins– PLD

Deactivation

• Proteolytic cleavage– Separation from regulatory domain– Degradation of function

• Phorbol ester mediated downregulation

PI-3 kinase

• Phosphorylates 3’ C of PI– PIP2PIP3

– p85 catalytic subunit, p110 regulatory subunit• RTKPI3KaktmTORprotein synthesis• RTKPI3Kakt--|FOXOapoptosis

Phospholipase D

• Cleaves PC to Phosphatidic acid+choline– PA

• Multiple activation factors– cPKC– ADP ribosylation factors (ARF)– Gangliosides GM1 & GM2

– RhoA

Phosphatidic Acid

• Convert to DAG, secondary PKC activation• Cofactor for PI-4P 5-kinase

– PIP2 synthesis– Raf/MAPK activation– mTOR

• Membrane biophysics– Negative charge, calcium attactor, pH drop– Budding– Vesicular transport

Phospholipase A2

• Cleaves sn-2 acyl chain from phospholipid• Calcium dependent cPLA2

– Specific for Arachidonic acid• Calcium independent iPLA2• Secreted sPLA2

– Not acyl chain specific– Membrane degradation

PLA2

• Arachidonic acid– Cyclooxygenase (COX) substrate– Prostaglandins, thromboxanes, prostacyclins– Paracrine factors

• Pro/anti-inflammatory• Vasoactive

• Phospholipid degradation– Venoms– Anti-coagulent– Atherosclerotic

Legend for following slide

PKC Active effector

DAGPLC

Second messenger

Enzyme which generates

PE20%

Primary phospholipid

Typical membrane content

Phospholipid metabolism

PI

PIP

PIP2

PIP3

DAGIP3

IP2

IP4

PA

MAG

IP

Inositol

PLC PLC

PI3K PTEN

DAG kinase

DAG lipase

8%

0.2%

0.2%

PI-5K

PI-4K

PA phosphatase

PKC

Ca2+ Raf kinasemTOR

PDK/mTOR

PC60%

PLD

Choline

LysoPCPLA2

PE

PS

20%

1%PSD

PEMT

CDP-DAGCDP-choline

Intercellular lipid signaling

Park et al 2012 Nat Rev Can 12:782

Angiogenesis

Inflammation

Prolific growth