Enzyme-coupled Receptors Cell-surface receptors 1. Ion-channel-coupled receptors 2. G-protein-coupled receptors 3. Enzyme-coupled receptors
Cell-surface receptors allow a flow of ions across the plasma membrane, which changes the membrane potential and produces an electrical current. Cell-surface receptors activate membrane-bound, trimeric GTP-binding proteins (G proteins, which then activate either an enzyme or an ion channel in the plasma membrane, initiating a cascade of other effects.
Cell-surface receptors either act as enzymes or associate with enzymes inside the cell; when stimulated the enzymes activate a variety of intracellular signaling pathways. Enzyme-coupled receptors Enzyme-Coupled Receptors 1. Activated RTKs recruit a complex of Intracellular signaling proteins 2. Most RTKs activate the monomeric GTPase Ras 3. RTKs activate PI 3-kinase to produce lipid docking sites in the plasma membrane 4. Some receptors activate a fast track to the nucleus 5. Multicellularity and cell communication evolved independently in plants and animals 6. Protein kinase network integrate information to control complex cell behaviors
Enzyme-coupled receptors Transmembrane proteins Ligand-binding domain on the outer surface Cytoplasmic domain acts as an enzyme itself or forms a complex with enzyme Discovered through growth factors that regulate the growth, proliferation, differentiation, and survival of cells Response slowly (hours) and require many intracellular transduction steps that eventually lead to changes in gene expression Direct, rapid responses (rapid reconfigulation of the cytoskeleton, controlling the way a cell changes its shape and moves) Fields of cancer biology and development (cell growth, proliferation, differentiation, survival, migration) Receptor tyrosine kinases (RTK), largest class of enzyme-linked receptors. Receptor tyrosine kinases (RTKs)
Some subfamilies of RTKs
1. Activated RTKs recruit a complex of intracellular signaling proteins Enzyme-coupled receptor An enzyme-linked receptor has to switch on the enzyme activity of its intracellular domain. Only one transmembrane segment (vs. GPCRs) It seems, no way to transmit a conformational change through a single α helix. A different strategy for transducing the extracellular signal.
Activation of an RTK 1. Binding of signal molecules 2. Receptor dimerization (Contact b/w the two receptor tails activate kinase function) 3. Cross-phosphorylation 4. Triggers the assembly of Intracellular signaling complex (The newly phosphorylated tyrosines serve as binding sites for a whole zoo of intracellular signaling proteins - 10 or 20 molecules) Proteins that become phosphorylated / activated. Adaptor - solely couples the receptor to other proteins 5. Assembled protein complex - trigger a complex response such as cell proliferation
1. 3-D structure of SH2 domain. The binding pocket for phosphotyrosine is shown in yellow on the right, and a pocket for binding a specific amino acid side chain is shown in yellow on the left. 2. The SH2 doamin is a compact, plug-in module, which can be inserted almost anywhere in a protein without disturbing the protein s folding or function. Termination of the activation of RTKs 1. Protein tyrosine phosphatase (removes the phosphates) 2. Endocytosis of the receptors and then destroyed by digestion in lysosomes
Different RTK recruits different intracellular proteins Common components 1.Phospholipase C (function in the same way as phospholipase C to activate the IP3 signaling pathway) 2.PI-3-kinase (Phosphatidyl-inositol 3-kinase): phosphorylates inositol phospholipids in the plasma membrane, which become docking sites for protein kinase B (PKB or Akt) 3.Ras (adaptor-assembled signaling complexes; a small GTP-binding protein) 2. Most RTKs activate the monomeric GTPas Ras
Ras protein Bound by a lipid tail to the cytoplasmic face of the plasma membrane All RTKs activate Ras (from PDGF to NGF) Small, monomeric GTP-binding protein (cf., trimeric G protein) Resembles the α subunits of a G protein Functions as a molecular switch Ras-GTP (active); Ras-GDP (inactive) Ras-activating protein (GEF or Sos); GTPase-activating protein (GAP) Activates a MAP kinase cascades Activation of Ras protein Grb-2 1. SH2 domain: phosphotyrosine of RTK 2. SH3 domain : proline-rich domain of GEF (Sos)
Ras Protein Ras activates a downstream serine/threonine phosphorylation cascade that include a MAP-Kinase
Raf Mek Erk MAP kinase signaling module : Ras > Raf > Mek-P > Erk-P > Gene regulatory proteins-p > control of gene expression Raf (MAP-kinse-kinse-kinase) MEK (MAP-kinase-kinase) Erk (MAP-kinase; mitogen-activated protein kinase) *mitogens: extracellular signal molecules that stimulate cell proliferation
1292 SnapShot: Ras Signaling Cell 133, June 27, 2008 2008 Elsevier Inc. Megan Cully and Julian Downward Cancer Research UK London Research Institute, London WC2A 3PX, UK DOI 10.1016/j.cell.2008.06.020 See online version for legend and references. The importance of Ras
The importance of Ras If the Ras is inhibited by an intracellular injection of Rasinactivating antibodies, a cell may no longer respond to some of the extracellular signals. If Ras activity is permanently switched on, the cell may act as if it is being bombarded continuously by proliferation-stimulating extracellular signals (mitogens). Ras in cancer cells (30% of human cancer) : a mutation in the gene for Ras causes the production of a hyperactive form of Ras. This mutant Ras helps stimulate the cells to divide even in the absence of mitogenes ( uncontrolled cell proliferation ). Oncogene and Proto-oncogene 2. Most RTKs activate the monomeric GTPas Ras
Enzyme-coupled receptors Enzyme-Coupled Receptors 1. Activated RTKs recruit a complex of Intracellular signaling proteins 2. Most RTKs activate the monomeric GTPase Ras 3. RTKs activate PI 3-kinase to produce lipid docking sites in the plasma membrane 4. Some receptors activate a fast track to the nucleus 5. Multicellularity and cell communication evolved independently in plants and animals 6. Protein kinase network integrate information to control complex cell behaviors 3. RTKs activate PI 3-kinase to produce lipid docking sites in the plasma membrane
Cell survival, growth, and proliferation RTKs Different RTK recruits different intracellular proteins Common components 1.Phospholipase C (function in the same way as phospholipase C to activate the IP3 signaling pathway) 2.PI-3-kinase (Phosphatidyl-inositol 3-kinase): phosphorylates inositol phospholipids in the plasma membrane, which become docking sites for protein kinase B (PKB or Akt) 3.Ras (adaptor-assembled signaling complexes; a small GTP-binding protein)
Review
PI 3-kinase-Akt Signaling Pathway PI 3-kinase (Phosphatidylinositol 3-kinase) Phosphorylates the inositol ring on carbon atom 3 to generate the inositol phospholipid. The two phosphorylated lipids serve as docking sites for signaling proteins with PH domains (Pleckstrin homology domain). PI 3-kinase-Akt Signaling Pathway Akt (protein kinase B or PKB) One of the relocated signaling proteins. Serine/threonine protein kinase Promotes the survival and growth of many cell types Inactivates the signaling proteins by phosphorylation Phosphorylate and inactivates a cytosolic protein, Bad Phosphorylation by Akt promotes cell survival by inactivating a protein that otherwise promotes cell death (apoptosis)
PI 3-kinase-Akt Signaling Pathway PDK1, mtor :Relocated PDK1 mtor PI 3-kinase-Akt Signaling Pathway
PI 3-kinase-Akt Signaling Pathway The PI 3-kinase-Akt Signaling Pathway Can Stimulate Cells to Survive Survival Signal > RTK-P > PI 3-kinase > 2x Inositol phospholipid-p > PDK1 (+ mtor) > Akt-P > BAD-P > Inhibition of Apoptosis *BAD: a protein that normally encourages cells to undergo programmed cell death, or apoptosis PI 3-kinase-Akt Signaling Pathway
PI 3-kinase-Akt Signaling Pathway The PI 3-kinase-Akt Signaling Pathway Also Can Stimulate Cells to Grow mtor mtor in complex 2: insensitive to rapamycin; it helps to activate Akt (cell survival). mtor in complex 1: sensitive to rapamycin; stimulates cell growth by promoting ribosome production and protein synthesis and by inhibiting protein degradation (cell growth). PI 3-kinase-Akt Signaling Pathway
3. RTKs activate PI 3-kinase to produce lipid docking sites in the plasma membrane 4. Some receptors activate a fast track to the nucleus
Not all enzyme-coupled receptors trigger complex signaling cascades that require a complex of intracellular kinases. Some enzyme-coupled receptors use a more direct route to control gene expression. A few hormones and cytokines (local mediators) ex., interferons (cytokines produced by infected cells that instruct other cells to produce proteins that make them more resistant to viral infection) STAT (for signal transducers and activators of transcription) - head straight for the nucleus, where they stimulate the transcription of specific genes. Jak-STAT signaling pathway Jak-STAT signaling pathway
Jak-STAT signaling pathway No intrinsic enzyme activity Instead, associated with cytoplasmic tyrosine kinase, JAK (Janus kinase) and STAT (Signal transducers and activators of transcription; Gene regulatory protein) JAKs STATs- (phosphorylate and activate) Migrate to the nucleus and stimulate transcription e.g., hormone prolactin, which stimulates breast cells to make milk, acts by binding to a receptor that is associated with JAKs. These JAKs activate STAT that turn on the transcription of genes encoding milk protein. Jak-STAT signaling pathway
Jak-STAT signaling pathway Cytokine/hormone binding Cross-links adjacent receptors Activates JAKs. Activated JAKs cross-phosphorylate one another and phosphorylate the receptor on tyrosine. STAT attaches to the phosphotyrosine on the receptor. JAK phosphorylates and activates STAT. STATs then dissociate from the receptor, dimerize, migrate to the nucleus. STATs activate the transcription of specific target genes. Hormone receptor > JAK- > STAT- (x2) > Tx Notch Signaling Pathway An even more direct signaling pathway Notch Controls the development of neural cells (remember Contact-dependent ) The receptor itself acts as a transcription regulator Binding of Delta Notch receptor is cleaved Releases the cytosolic tail of the receptor Heads to the nucleus and activate transcription The simplest and most direct way Delta binding > Notch cleavage > Tx
Contact-dependent signaling controls nerve-cell production Notch Signaling Pathway
4. Some receptors activate a fast track to the nucleus 5. Multicellularity and cell communication evolved independently in plants and animals
Multicellularity in Plants and Animals Evolved independently ( every cell for itself ; evolved its own molecular solution to the complex problem of becoming multicellular; evolved separately) Some similarities Transmembrane cell-surface receptors (e.g., Arabidopsis thaliana ( ) has genes for receptor serine/ threonine kinases) However, not to use RTKs, steroid-hormonetype nuclear receptors, or camp, and few GPCRs.
Ethylene signaling pathway in plants One of the best-studied signaling systems in plants. Ethylene receptor Ethylene, a gaseous hormone that regulates a diverse array of developmental processes. Not belong to any of the classes Dimeric transmembrane proteins. Ethylene (-) Empty receptor activates a protein kinase Transcription regulator degraded and the ethylene responsive genes shut off. Ethylene (+) Receptor and kinase are inactive ethylene-responsive genes are transcribed. This strategy (signals to relieve transcriptional inhibition) is common in plants. 5. Multicellularity and cell communication evolved independently in plants and animals
6. Protein kinase networks integrate information to control complex cell behaviors
Complex Signaling Pathways Complexity of cell signaling is much greater than we have described. We have not discussed every intracellular signaling pathway available to cells. cross-talk (the kinases often phosphorylate components in other signaling pathways) About 2% (400 genes) of our ~20,000 protein coding genes in our genome code for protein kinases. Hundreds of distinct types of protein kinase in a single cell. A tangled web 1292 SnapShot: Ras Signaling Cell 133, June 27, 2008 2008 Elsevier Inc. Megan Cully and Julian Downward Cancer Research UK London Research Institute, London WC2A 3PX, UK DOI 10.1016/j.cell.2008.06.020 See online version for legend and references.
What is the function of such complexity?
A cell receives messages from many sources, and it must integrate this information to generate an appropriate response: to live or die, to divide or differentiate, to change shape, to move, to send out a chemical message of its own, and so on. Integrating protein (protein kinases) Usually have several potential phosphorylation sites, each of which can be phosphorylated by a different protein kinases. Information received from different sources can converge on such proteins In turn, can deliver a signal to many downstream targets.
But, our understanding of these intricate networks is still evolving: we are still discovering new links in the chains, new signaling partners, new connections, and even new pathways.
6. Protein kinase networks integrate information to control complex cell behaviors