Biol 2056 2024 L1 Signal Transduction PDF

ome concepts of signal transduction Biol 2056 2024 L1 Membrane translocation is a common event in signal transduction Biol 2056 processes 2024 L1 Volume=4000 um3 Surface area=1200um2 Consider that after t...

ome concepts of signal transduction Biol 2056 2024 L1 Membrane translocation is a common event in signal transduction Biol 2056 processes 2024 L1 Volume=4000 um3 Surface area=1200um2 Consider that after translocation to the Membrane the protein is within the 5nm of the membrane (approx. diameter of a protein) then these proteins experience a volume of 1200X0.005=6um3.that is a 700fold concentration! Binding and enzymatic reactions are dependent on the T shows that proteins tethered to the membrane interact more. Biol 2056 2024 L1 Two non interacting proteins Two non interacting proteins in the cytosol Targeted to the membrane with lipid tether 480nm 560nm 480nm 560nm GFP RFP GFP RFP Strong FRET signal No FRET signal Fluorescent resonance energy transfer: depends on the distance between fluroph ff with the distance. Must be with approximately 10nm (ie interacting) ced Membrane localisation of PKB drives cell transformation Biol 2056 2024 L1 echanisms to control protein localisation at the membrane Biol 2056 2024 L1 Protein interaction Lipid interaction motif Lipid tether SH2 P RAS P P KB e as Ki n 1. Phosphobinding 1. Phosphoinositide 1. Myristoylation motifs: SH2, PTB interacting motifs: PH, 2. Prenylation 2. Ubiquitin binding FYVE, PX, PHD and lysine Often single lipid motifs: arginine rich patches. tether not enough 3. AKAP interaction 2. DAG binding motifs: C2 for stable membrane domains (interaction domain localisation and the between a cytosolic 3. Membrane interacting tether can be and a membrane motifs: C1 domain, combined with lipid  Biol 2056 yristoylation : controlling membrane localisation 2024 L1 1. Fatty acylation: mainly consists of the covalent addition of palmitic or myristic fatty acids to proteins. 2. covalent addition of the 14-carbon saturated fatty acid myristate to the N-terminal glycine residue through a stable amide bond. 3. Thought to be irreversible. A. Removal of the N-terminal methionine B. Activation of myristic acid with CoA C. Coupling of the myristic acid to the glycine. D. Gly-X3-X4-X5-(Ser/Thr/Cys)6 where X represents most amino acids, except for proline, aromatic or charged residues in position X3 E. This is co-translational myristoylation.  Biol 2056 st-translational myristoylation: role in apoptosis. 2024 L1 Caspase cleavage. 1. Caspase mediated cleavage of Bid exposes a glycine residue 2. Bid becomes myristoylated 3. Lipid tether induces insertion into mitochondrial membrane 4. Recruitment BAK to the mitochondrial membrane 5. Cytochrome C release 6. Downstream apoptosis.  Biol 2056 Protein prenylation 2024 L1 1. Mainly occurs on CAAX proteins : Proteins containing a CAAX motif at their carboxyl termini, in which ‘C’ is the Cys residue that functions as the isoprenoid attachment site, ‘A’ signifies any aliphatic amino acid, and ‘X’ denotes any of several amino acids. Generates a thioether linkage. 2. prenylation is initiated by the attachment of a 15‑carbon (farnesyl) or a 20‑carbon (geranylgeranyl) isoprenoid lipid to the Cys residue by protein farnesyltransferase (FTase) or protein geranlygeranyltransferase I (GGTase I), respectively. 3. Proteins can then be further processed by RAS-converting CAAX endopeptidase 1 (RCE1), which removes the -AAX residues. 4. isoprenylcysteine carboxylmethyltransferase (ICMT), ‘caps’ the carboxyl group on the now carboxy‑terminal isoprenoid-modified Cys residue with a methyl group  Biol 2056 ylation and palmitoylation of RAS controls its membrane localisation 2024 L1 15 carbon farnesyl diphosphate (FPP) 20‑carbon geranylgeranyl diphosphate (GGPP) Both built from isopentenyl diphosphate (IPP). Prenyl group put on by farnesyl transferase or geranylgeranyltransfe rase. Ft-inhibitors developed for the clinic, however RAS instead become modified with geranylgeranyl. Statins used in clinic to control cholesterol levels. Metadata suggest that long- term statin therapy  Biol 2056 gnal regulated membrane localisation 2024 L1 1. Single Lipid tethers not normally enough to induce stable membrane localisation. 2. Often requires other membrane binding domains. A. Phosphoinositide interaction domain. Often polybasic regions. B. Another tether such as a prenylation or palmitoylation 3. As the tether does not induce stable association this enables various control mechanisms to be used. ---------------------------- ---------------------------- +++++ MARCK S +++++ MARCK recoverin S Polybasic region GTP binding Ca2+ binding Phosphorylation Conformational Conformational with the PBR switch switch induces membrane dissociation. Kras4B is targeted to the PM by the interaction of its polybasic Biol 2056 region with phosphoinositides 2024 L1 1. Ras mutated in 16% of human tumours 2. Mutation in codon 12, 13 and 61 3. Kras is most highly mutated H and N Ras undergo prenylation and palmitoylation to target them stably to the membrane. Kras on the other had only undergoes prenylation. It uses a Serine 181 is a target for PKC PBR for stable association at the Mediated phosphorylation  Biol 2056 ospho-switch controls Kras localisation at the PM and downstream functions. 2024 L1 GOLGI mitochondria PKC induced Bryostatin Apoptosis is (PKC activator ) R GTP effector ?? Apoptosis I dependent Induces a P farnesylation Switch ation of the farnesylation switch attenuates tumour growth in vitro and in vivo progeria syndrome such as Hutchinson–Gilford (HGPS), are caused Biol 2056 by abnormal processing of the CAAX protein prelamin A 2024 L1  Biol 2056 se inhibitors can be used in the clinic to treat HGPS 2024 L1 e lamins maintain isoprenylation pe lamins are cleaved as below pe lamins are produce as mature proteins without caax or clevage. Abnormal proteolytic cleavage of lamin A leads to HGPS. Loss of the function of ZMPSTE24 Mutation of the cleavage site in Lamin A Both lead to an accumulation of prenylated lamin Treatment with Ftase inhibitors shows promise in clinical  Biol 2056 questration can be used to inhibit downstream signalling 2024 L1 Notch Organelle specific sequestration 1. Plasma ER membrane: TUBBY proteins and Notch 2. ER. Unfolded protein response :CHOP 3. Lysosome: TFEB 4. Cytoplasm : steroid  Biol 2056 n phosphorylation is a common event in signal transduction pathways 2024 L1 ding strongly suggest you read and digest: he regulation of protein phosphorylation (Johnson). Cell Signaling by Receptor Tyrosine Kinases. (Lemmon and Joseph Schlessinger) Tyrosine Hippo GPCR Kinase cytokine Ion channels steroids WNT Receptor recepto rs Transduction Protein phosphorylation Pathway Differential output signalling and cell fate determination output The regulation of Protein phosphorylation is a major pathway Biol 2056 downstream of receptor activation that mediates cell specific 2024 L1 output signalling. Writer: Protein Kinase Protein Protein Specific X X cellular output Erazer: Protein phosphatase  Biol 2056 osphorylation as a direct regulator of protein conformation 2024 L1 P Conformational change Phosphorylation can change conformation by either disrupting molecular interactions or by promoting them. Common event in protein kinase activation. Phosphorylation creates a new binding site for a phospho- Biol 2056 specific reader molecule that delivers the output 2024 L1 Readers Writer: Protein Kinase Tyrosine P Read Protein Protein er Specific X X cellular output threonine P Erazer: Protein phosphatase Serine/threonineP Phosphorylation readers such as PIN1 can induce conformational Biol 2056 changes 2024 L1 Writer: Proline directed Kinase MAPK, p38 stress kinase Cyclin kinases Catalytic activity Protein Protein Interaction partners Pin1 X X Subcellular localisation Erazer: Dephosphorylation Protein phosphatase Pin1= peptidyl-prolyl cis/trans isomerase. Only isomerises a proline near to a phosphor_threonine or serine  Protein phosphorylation induces diverse downstream outputs Biol 2056 2024 L1 P Conformational change Intramolecu lar Regulation Tyrosine P Induce or eliminate Protein interaction Localisati on threonine P (FOXO and 14-3- 3, KRAS Stability PBR) (EGFR Serine/threonineP and CBL) Downstream signalling cascades A multitude of post-translational protein modification is used to Biol 2056 drive the regulation of downstream pathway signalling. 2024 L1 Ub writer M reader output Eraser  Biol 2056 Writer/Reader modules can be coupled to induce complex outputs 2024 L1 Remember cbl and the tyrosine kinase receptors.  PTMS; what are they good for? Its all a matter of states! Biol 2056 2024 L1 PTM of the P53 tumour suppressor P53 tumour suppressor : mutated in over 50% of human cancers. Pathway probably mutated in all cancers