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Transcript
BCOR 011 Cell Communication II Lect 19 Signal molecule Activated Ras-GTP A G-Protein Receptor Chapt 11 Inactive MAPKKK 1 Active MAPKKK 1 Inactive MAPKK 2 ATP Protein Phosphatases P ADP Pi 10/17/05 And they tell 2 friends And they tell 2 friends And they tell 2 friends… Active MAPKK 2 PP Inactive MAPK 3 ATP P ADP Pi Active MAPK 3 PP Inactive protein ATP P ADP Pi PP Active protein Cellular 1response Lecture Outline 1. Finish Trimeric G-Protein: Phospholipase C -DAG, IP3 second messengers Protein Kinase C, Ca++ release 2. Nitric Oxide, cGMP activation - consequences of failing to inactivate 3. Tyrosine Kinase Receptor - RAS G-protein, MAP Kinase Cascade - Cell Cycle Control, Gene Control 4. Internal Receptors – Steroid Receptor - Gene Regulation 2 TWO subclasses of trimeric G-protein-activated signal transduction pathways: A. target protein adenylate cyclase cAMP-> PKA B. target protein phospholipase C 3 target effector enzyme is Phospholipase C PLC cleaves a membrane phospholipid (Phoshatidyl inositol) to two 2nd Messengers: Inositol-1,4,5-Trisphosphate (InsP3) & Diacylglycerol (DAG)4 PIP2 DAG Lipid Soluble InsP3 Water Soluble 5 DAG Activates Protein Kinase C (Starts Cascade) InsP3 Ligand for ER ligandgated Ca++ channels 6 Ca++ levels 1 A signal molecule binds to a receptor, leading to activation of phospholipase C. EXTRACELLULAR FLUID 2 Phospholipase C cleaves a 3 DAG functions as a second messenger in other pathways. plasma membrane phospholipid called PIP2 into DAG and IP3. Signal molecule (first messenger) G protein DAG GTP PIP2 G-protein-linked receptor Phospholipase C IP3 (second messenger) IP3-gated calcium channel Endoplasmic reticulum (ER) Various proteins activated Ca2+ Figure 11.12 Cellular response Ca2+ (second messenger) 4 IP quickly diffuses through 3 the cytosol and binds to an IP3– gated calcium channel in the ER membrane, causing it to open. 5 Calcium ions flow out of 6 The calcium ions the ER (down their concentration gradient), raising the Ca2+ level in the cytosol. activate the next protein in one or more signaling pathways. 7 Response: Protein Kinase C phosphorylates target proteins (ser & thr) cell growth regulation of ion channels cytoskeleton increases cell pH Protein secretion Ca++ Binds & activates calmodulin Calmodulin-binding proteins activated (kinases & phosphatases) 8 Shut Off - remember, signal needs to be transient - Must shut off cascade: removal of ligand, (self)-hydrolysis of GTP, remove IP3, protein phosphatases, Ca++ ion pumps 9 Direct activation signal: nitric oxide (NO) It’s a gas! lipid soluble, binds directly to activate enzyme made by: endothelial cells (line blood vessels) target: guanylyl cyclase GTP->cGMP NO synthase response: relaxes smooth muscle vessels dilate, blood flow +NO arginine citrulline 10 target Shut off by cGMP phosphodiesterase 11 Nitroglycerine – taken to relieve angina 1977 –Ferid Murad Nitroglycerine acts to elict release of NO, relaxes cardiac muscle 1998 Nobel Prize – Murad, Furchgott, Ignarro 12 Nitro glycerine Shut off By cGMP phosphodiesterase 13 Sildenafil citrate (VIAGRA) 14 cell-surface receptors a. ion-channel-linked b. G-protein-linked c. Protein-kinase associated (enzyme-linked) 15 Mitogen Activated Protein-Kinase associated receptors receptor has enzymatic activity - only when ligand binds ligand: allosteric effector -> conformational change receptor tyrosine kinases Growth & differentiation control of cell cycle control of gene expression bad news when messed up - cancer 16 Growth factor Reception Receptor Growth Factor “Mitogen-activated” Phosphorylation cascade Transduction Signaling Cascade CYTOPLASM Inactive transcription factor Active transcription factor Response P DNA Gene NUCLEUS Figure 11.14 mRNA 17 EGFR – epidermal growth factor receptor •Monomer receptor 1 transmembrane segment •Ligand binds – receptors dimerize •Activates Tyr kinase •autophosphorylate •Scaffold to Bind/activate Target proteins •GDP/GTP exchange •Activate Ras G-protein EGF – growth factor or mitogen 18 “Mitosis-generator” Receptor tyrosine kinases Signal-binding site Signal molecule Signal molecule Helix in the Membrane Tyrosines Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Tyr Receptor tyrosine kinase proteins (inactive monomers) CYTOPLASM Dimer Figure 11.7 Activated relay proteins Tyr Tyr P Tyr Tyr P P Tyr Tyr P Tyr Tyr P Tyr Tyr P P Tyr Tyr P Tyr Tyr P Tyr Tyr P P Tyr Tyr P 6 Activated tyrosinekinase regions (unphosphorylated dimer) ATP 6 ADP Fully activated receptor tyrosine-kinase (phosphorylated dimer) Inactive relay proteins Cellular response 1 Cellular response 2 19 Receptor tyrosine kinases can activate ras ras is a monomeric G-protein “molecular switch” 20 A phosphorylation cascade Signal molecule Ras-GTP Receptor MAP Kinase Inactive protein kinase Kinase 1 Kinase Activated relay molecule MAP Kinase Active protein Kinase kinase 1 Kinase MAP Inactive Kinase protein kinase 2 Kinase phosphatases (PP) catalyze the removal of the phosphate groups from the proteins, making them inactive and available for reuse. 2 Active protein kinase 1 transfers a phosphate from ATP to an inactive molecule of protein kinase 2, thus activating this second kinase. P MAP Active protein Kinase kinase 2 Kinase ATP ADP Pi 5 Enzymes called protein 1 A relay molecule activates protein kinase 1. PP MAP Kinase Inactive protein kinase 3 ATP Active MAP protein kinase Kinase 3 ADP Pi 3 Active protein kinase 2 then catalyzes the phosphorylation (and activation) of protein kinase 3. PP Inactive protein P 4 Finally, active protein kinase 3 phosphorylates a protein (pink) that brings about the cell’s response to the signal. ATP P ADP Figure 11.8 Pi PP Active protein Cellular 21response Ras activation sets off a phosphorylation cascade 10 100 MAPKK 1,000 MAPK 100,000 MAPKKK Mitogen Activated Protein Kinases MAPKs Controls: -Transcription Factors -Translation Factors 22 -Cell Division How do you turn it off? phosphatases GTPase (GTP->GDP + P) molecular switch on internal timer If timer broken – on all the time 23 PROBLEMS IN CANCER: - broken ras – won’t shut off - Broken receptor – thinks ligand there even when it isn’t -broken MAPK – on all the time, even when not phosphorylated RESULT: continuous signal for cell to divide 24 Specific signal transduction cascades: 1. receptor-mediated cell-surface 2.receptor-mediated intracellular Lipid soluble things: steroid hormones 25 Steroid Hormone Plasma membrane Lipid soluble Crosses membranes Receptor protein Binds intracellular receptor In cytosol steroid receptor Complex Changes shape Releases from tether protein Travels to Nucleus steroid receptor Binds DNA Turns genes ON EXTRACELLULAR FLUID Hormone (testosterone) Testosterone binds to a receptor protein in the cytoplasm, activating it. The hormonereceptor complex enters the nucleus and binds to specific genes. Hormonereceptor complex The bound protein stimulates the DNA transcription of the gene into mRNA. mRNA NUCLEUS Figure 11.6 The steroid hormone testosterone passes through the plasma membrane. The mRNA is translated into a specific protein. New protein 26 CYTOPLASM The Specificity of Cell Signaling • The different combinations of proteins in a cell – Give the cell great specificity in both the signals it detects and the responses it carries out Same hormone can give different responses in different cells 27 Pathway “cross-talk” Signal molecule Receptor Relay molecules Response 1 Cell A. Pathway leads to a single response Response Response 2 3 Cell B. Pathway branches, leading to two responses Figure 11.15 Activation or inhibition Response 4 Cell C. Cross-talk occurs between two pathways Response 5 28 Cell D. Different receptor leads to a different response Specific pathways 1. Cell-surface receptor mediated a. ion-channel-linked b. trimeric G-protein-linked (i) adenylyl cyclase cAMP ->protein kinase A (ii) phospholipase C InsP3, DAG, Ca++, protein kinase C Direct activation - NO c. protein kinase-associated (enzyme-linked) Receptor tyrosine kinase monomeric G-protein (ras), MAPKs 29 2. Intracellular receptors – steroid hormones, dioxin