Quiz 1 Material Lipids Membranes Permeability Transport Channels Membrane Poten9al Receptors as Channels GPCRs RTKs Phosphoinosi9des PIPs Generic GOProtein Signaling Ac4va4on Inac4va4on 1 GLprotein w GTPL separates from 2 Targets ac9vated 3 G LGTP hydrolyzes its own GTP to GDP 4 G reLbinds GLprotein returns to res9ng state GOProtein Signaling Adrenalin Pathway signal s9mulus adrenalin receptor Ladrenergic transducer Gs 2 receptor protein target adenylyl cyclase 2nd messenger increase in cAMP result ac9ve Protein Kinase A PKA Turning O OAdrenergic Signaling 1 Adrenalin released from receptor 2 GAP s9mulates GTPase by Gs 3 GsL hydrolyzes GTP 4 Phosphodiesterase cleaves cAMP 5 PKA turns o as regulatory cataly9c subunits assemble 6 Phosphoprotein phosphatase turns o PKA targets 7 BARK phosphorylates receptor in ac9ve state 8 Larres9n binds phosphorylated receptor to STOP SIGNAL OAdrenergic Receptor Kinase BARK GPCR kinase GRK of Ladrenergic adrenalin receptor func9ons to turn o adrenalin signaling by phosphoryla9ng cytoplasmic side of Ladrenergic receptor ac9ve adrenalin receptor s9mulates BARK to phosphorylate receptor when receptor is adrenergic receptor kinase BARK phosphorylated Larres9n recognizes binds receptor to turn o signaling New findings also implicate arrestin in other functions Promote endocytosis Activation of MAP kinase RTK by cross talk Scaffold protein to organize signaling pathways Lefkowitz and Shenoy Science 308 512 517 2005 Retina Adenylyl Cyclase enzyme in adipocyte GLprotein signaling ac9vated by GsL turns ATP into cAMP 2nd messenger in Ladrenergic signaling pathway Protein Kinase A PKA result of Ladrenergic signaling pathway PKA ac9vated by cAMP PKA pathway regulates metabolism gene is ac9vated expression GOProtein Signaling In the Re4na signal s9mulus photon of light receptor rhodopsin transducer transducin target cGMP phosphodiesterase cleaves cGMP 2nd messenger decrease in cGMP i e ligand for ligandLgated Na channel result close Na channel GOProtein Signaling Light Closes Ca4on Channels in the Re4na Light Closes Cation Channels in the Retina Na channel OPEN in DARK depolarized Na channel CLOSED in LIGHT repolarized inhibitory signals released in dark but not in light cGMP Gated Channel Receptor Tyrosine Kinase RTK Ac4va4on 1 Bind ligand 2 Dimerize 3 AutoLphosphoryla9on on tyrosine 4 Ac9vate via transLphosphoryla9on Ac4vated RTKs Assemble Large Mul4OMolecular Complexes Activated RTK cid 1 s Assemble Large Multimolecular Complexes SH2 SRC Homology 2 binds phosphorylated tyrosine Protein Motifs Src Homology 2 SH2 Binds PTB Protein Tyrosine phosphorylated Binding binds tyrosine Protein Tyrosine phosphorylated tyrosine Binding PTB SH3 SRC Homology 3 Binds binds prolineLrich regions PH Pleckstrin Homology phosphorylated tyrosine SRC Homology 3 SH3 Bind proline rich domain binds phosphorylated inositol lipid Pleckstrin Homology regions PH Domain Bind phosphorylated inositol lipid SH2 Pathway to Ras Pathway to Ras adaptor Grb2 GEF SOS couple RTK to Ras Grb2 has 1 SH2 binds RTK Grb2 has 2 SH3 s bind SOS Ras has a GTPase cycle RasLGEF ac9vates Ras RasLGAP inac9vates Ras ac9vated RTKs control RasLGEF RasLGAP Adapter Grb2 and GEF SOS Couple RTK to Ras Grb2 has one SH2 binds RTK and two SH3 bind SOS Ras Ras Structure switch regions do not bind GDP are displaced by SOS do contact GTPL LP decrease GTPase maintain Ras in ac9ve state MAP Kinase Cascade MAP Kinase Cascade Three Protein Kinases inac9ve RAF is bound to 2 mol MAP KKK RAF 14O3O3 MAP KK MEK a S T Y kinase i e phosphorylates substrates on S T Y MAP K ac9va9on lip has T183 Y185 ac9ve form leads to ac9va9on of transcrip9on signals induce shortLterm longLterm gene expression responses MAP KKK RAF MAP KK MEK MAP K ERK Inactive RAF is bound to 2 moles 14 3 3 Ras GTP Displaces 14 3 3 and activates RAF MEK is a S T and Y Kinase Note MAP K signals induce short term and long term gene expression responses MAP K Overview of Signal Transduc4on Pathways Overview of Signal Transduction Pathways Phosphoinosi4de Signaling 1 Ac9vate Phospholipase C PLC PLC clips PIP2 to DAG IP3 PLCL ac9vated by GPCRs PLCL ac9vated in RTK pathway 2 IP3 releases Ca2 IP3 important in Ca2 Ldependent signaling i e phosphoinosi9de signaling b c it can travel a much longer distance in cytosol than Ca2 can IP3 travels to ER lumen binds IP3Lgated Ca2 Ldependent release channel Ca2 travels shorter distance to ac9vate PKC DAG Ca2 ac9vate PKC Promotes cell growth Ac9vate Phospholipase A2 PLA2 Ac9vate PIO3K PhosphoinositolL3 Kinase via RTK pathway PH domain recruited PIO3K phosphorylates PIP2 to PIP3 on cell membrane PKB Akt binds PIP3 Ac9vate PKB Akt w PDK1 mTOR growth 3 4 5 6 7 8 9 10 Di eren9a9on occurs 11 Ac9ve PKB Akt phosphorylates BAD to release death inhibitory protein i e block programmed cell death Targets of Ca2 in Cytoplasm Calmodulin dumbbell structure 4 EF hands bind Ca2 calmodulin folds around target hydrophobic interac9on CAM Kinase II ac9vated in the presence of Ca2 calmodulin Myosin Light Chain Kinase Protein Kinase C PKC tumor promoter in presence of mutagen makes mutagen work be er DAG PS Phospha9dyl Serine lipids that contribute to ac9va9on substrates ac9vated downstream Na H an9port 5LP tase cleaves phosphate from C5 of IP3 opens Ca2 channel to produce IP2 does not open Ca2 channel stops Ca2 signaling in summary turn on PKC Ca2 signaling is limited PIO3K PI 3 Kinase a kinase that phosphorylates a phospholipid on C3 of inositol ring ac9va9on required for growth factors to promote cell division ac9va9on can be promoted either via RTK pathway or by Ras promotes cell division PTEN signal turned o by 3Lphosphatase tumor suppressor PI 3 Kinase promotes cell division 3 Phosphatase turns off the Signal PTEN PTEN is a tumor suppressor PKB Protein Kinase B PKB Akt requires PH domain to bind 3Lphosphoinosi9des phosphoryla9on of PKB by PDK1 PDK1 a kinase w its own PH domain Activation of Protein Kinase B Akt Requires Binding of the PH Domain to 3 phosphoinositides and Phosphorylation of PKB by PDK1 Note PDK1 is a protein kinase that has its own PH domain PKB Regulates Apoptosis PKB Ac4vates mTOR to Promote Cell Growth PI 3K Pathway Promotes Growth Survival Protein Kinase B Activates mTOR to Promote Cell Growth mTOR is mammalian Target Of Rapamycin protein kinase growth factors bind RTKs s9mulate protein synthesis ac9vate PIO3K ac9vate PKB Akt inac9vate TscO2 GAP that inac9vates Rheb RhebLGTP ac9vates mTOR mTOR promotes cell growth by s9mula9ng