Unit 8 Flashcards
(12 cards)
G-Protein coupled receptors also signal through phospholipids & Ca2+
1.) Ligand binding activates Gqα, separates it from the βγ subunits
2.) Gqα activates PLC-β, leading to the release of DAG & IP3
3.) IP3 diffuses through the cytosol to Ca2+ channels on the ER
4.) The Ca2+ channels open, releasing Ca2+ into the cytosol
5.) The rise in intracellular Ca2+ allows Ca2+ to bind to protein Kinase C (PKC) causing the protein to relocate to the plasma membrane
6.) PKC at the plasma membrane associates with, and is activated by, DAG
7.) Activated PKC can then phosphorylate the numerous downstream targets to elicit different cellular responses
Glucose in the blood is a signal for the release of insulin from pancreatic cells
1.) As glucose rises above 5 mM, it is imported through the GLUT2 transporter
2.) This leads to an increase in glycolysis, producing ATP & pyruvate
3.) ATP binds to ATP-sensitive K+ channels, closing the channel
4.) The closed channels lead to a slight depolarisation of the plasma membrane
5.) Depolarisation causes the opening of voltage-sensitive Ca2+ channels
6.) The increase in intracellular Ca2+ triggers the release of secratory vesicles containing insulin
Regulation of muscle contraction by Ca2+ and CaM
1.) A rise in intracellular Ca2+ causes Ca2+ to bind to CaM
2.) CaM Ca2+ activates CaMKII Ca(aM kinase)
3.) CaMKII enters the nucleus through the pores & phosphorylates CREB
4.) CREB dimerises & binds to DNA at CRE sequences & recruits p300 & CBP to activate gene transcription
Regulation of CaMP levels by Ca2+ and CaM
1.) A rise in intracellular Ca2+ causes Ca2+ to bind to CaM
2.) CaMCa2+ activates calcium-calmodulin dependent phosphodiesterase 1 (PDE1), highly expressed in brain, heart & lung
3.) PDE1 converts cAMP (produced by adenylyl cyclase) to AMP, downregulating cAMP-dependent signaling pathways
The Ras-Raf-MAP kinase pathway
- Ras-small GTP binding protein. When bound to GDP it is inactive, when bound to GTP it is active
- GRB2: An SH2 domain-containing protein (binds to phosphotyrosine residues). Mediates attachment of Sos to the activated EGF receptor
- Sos-Nucleotide exchange factor for Ras, activates the GTPase
Overview of the Ras-Raf-MAP kinase pathway
- Analogous pathways activated by different receptors exist
- The three kinases (Raf, MEk, & ERKII / 2 can be generally classified as follows:
1.) The most downstream kinase of the three (ERK 1/2) is called the MAP kinase (MAPK)
2.) Since MEK is the kinase that phosphorylates the MAPK, it is called a MAPK Kinase (MAPKK)
3.) Since Raf is the kinase that phosphorylates the MAPKK, it is called a MAPKK kinase (MAPKKK) - All similar pathways conform to this kinase structure
Mechanisms of silencing the cascade
1.) GTP hydrolysis by Ras (mutation of Ras to a form that cannot hydrolyse GTP is a common mutation found in cancer cells)
2.) Dephosphorylation of the MAPK (one gene whose transcription is upregulated by ERK1 is MKP-1, a phosphatase that acts on ERK1)
3.) Inactivation of the receptor by:
a.) receptor dephosphorylation
b.) Endocytosis & delivery to the lysosome for degradation
c.) Therapeutic method - antibody inhibition. Herceptin is an antibody that recognises Her2 (aka EGF receptor) and is used to treat breast cancer.
Signaling through proteolysis
Notch-Delta pathway. Important for cell fate determination
1.) Delta acts as the ligand & Notch as the receptor. Both are transmembrane proteins
2, 3.) When the 2 interact, Notch is cleaved by 2 proteases
a.) ADAM1O - cleaves extracellularly
b.) Presenilin1 - cleaves within the membrane-spanning region & releases a fragment of the Notch intracellular domain
4.) This domain then translocates to the nucleus where it activates transcription
Convergence
- When receptors can activate similar pathways. Thus, different signals may lend to expression of similar genes. For example, 2 different receptors may lend to the recruitment of GRB2-Sos, activating Ras-Raf-MapK pathway
Divergence
- When a single receptor can activate numerous pathways. For example the insulin receptor activates PI38, GRB2-Sos and other pathways
Cross-talk
- When part of one pathway influences part of another pathway. For example, activated PKA phosphorylates & inhibits the function of Raf, thereby blocking Raf signaling pathways
Methods of studying a signaling pathway
1.) Protein-protein interaction methods
2.) Mutagenesis to determine the key amino acids required for interaction & for signaling
3.) Using bypass experiments with mutant cell lines to determine the order of the pathway
