Cell Signaling and Signal Transduction PDF

CHAPTER 15 Cell Signaling and Signal Transduction: Communication Between Cells Required reading: relevant sections of Chapter 15 15.1-1  15.1 | The Basic Elements of Cell Signaling Systems Extracellular signal to cellula...

CHAPTER 15 Cell Signaling and Signal Transduction: Communication Between Cells Required reading: relevant sections of Chapter 15 15.1-1  15.1 | The Basic Elements of Cell Signaling Systems Extracellular signal to cellular response – general principles 1: synthesis of the signaling molecule 2: release of the signaling molecule via exocytosis 3: transit of signaling molecule to the target cell 4: binding of signaling molecule (ligand) to a protein receptor on the target cell 5: binding of ligand to receptor results in a conformational change of the receptor 6: receptor initiates one or more intracellular pathways that results in changes in: cellular function Metabolism gene expression Shape 7: deactivation of the receptor movement 8: removal of ligand 15.1-2  15.1 | The Basic Elements of Cell Signaling Systems Types of intercellular signaling: transmission between cells a) endocrine signaling, messenger molecules reach their target cells through the bloodstream (example = insulin) b) paracrine signaling, messenger molecules travel short distances through extracellular space example = neurotransmitters c) autocrine signaling, the cell has receptors on its surface that respond to the messenger 9example = T-cells during an immune response d) Juxtacrine signaling (or contact dependent signaling): short range but requires physical contact between sending and receiving cells (example = antigen presentation) 15.1-3 15.1 | The Basic Elements of Cell Signaling Systems: Receptor location 1. Cell-surface receptors Most signaling molecules are hydrophilic and are unable to cross the plasma membrane: They bind to cell surface receptors Three main classes of cell surface receptors: G-protein coupled (GPCR) Enzyme-linked Ion Channel-linked 2. Intracellular receptors Small hydrophobic signaling molecules can diffuse across the plasma membrane and bind to receptor proteins either in the cytoplasm or nucleus 15.1-4 15.1 | The Basic Elements of Cell Signaling Systems Two different types of signaling pathways are shown in this figure Receptors on or in target cells receive the message Some cell surface receptors generate a soluble and diffusible intracellular second messenger Second messengers are small substances that activate (or inactivate) specific proteins Other surface receptors recruit proteins to their intracellular domains at the plasma membrane Most signal transduction pathways use a combination of both mechanisms 15.1-5 15.3 | The Basic Elements of Cell Signaling Systems First messenger = ligand Second messenger = small molecules that increase or decrease in concentration in response to first messenger (can be anywhere along the cascade) Second messengers bind to other proteins to modify their activity 15.1-6 15.1 | The Basic Elements of Cell Signaling Systems: protein kinses and phosphatases  15.1 | The Basic Elements of Cell Signaling Systems Molecular switches: phosphorylation The addition of phosphate groups to hydroxyl groups on (most commonly) serine threonine tyrosine Kinases phosphorylate Phosphatases dephosphorylate Phosphorylation changes a proteins charge and generally leads to a conformation change which can alter ligand binding or other features of the protein resulting in an increase OR decrease of its activity Phosphorylation is part of (almost) all signaling pathways Human genome contains 600 protein kinases 100 protein phosphatases 15:11  15.1 | The Basic Elements of Cell Signaling Systems Molecular switches: GTP-binding proteins GTPase superfamily: enzymes that hydrolyze GTP to GDP Two conformations on = bound GTP that modulates the activity of specific target proteins to which they bind off = bound GDP GAPS: GTPase-activating proteins RGSs: regulators of G protein signaling Remember that GDI: guanine nucleotide dissociation GDP is exchanged inhibitors for GTP GEFs: guanine nucleotide exchange factors 15:13 15.1 | The Basic Elements of Cell Signaling Systems A comparison in the frequency of tyrosine phosphorylation (by red intensity) in two different types of breast cancer cells Triple-negative cells do not express the three major molecular signatures of breast cancer cells- estrogen receptor, progesterone receptor, and the growth factor HER2 triple-negative cells have a much greater level of tyrosine phosphorylation than other breast cancer cells This may correlate with the loss of a particular protein tyrosine phosphatase activity (PTPN12) in many of the triple- negative cancers 15:12 15.1 | The Basic Elements of Cell Signaling Systems Signals are often amplified: A small amount of ligand can illicit a large response from a target cell Remember that protein interaction alters conformation: Often occurs by phosphorylation or dephosphorylation 15:10 15.1 | The Basic Elements of Cell Signaling Systems Many different ways in which signals can be integrated Remember that protein interaction alters conformation 15:8  15.1 | The Basic Elements of Cell Signaling Systems Many (most) signaling proteins are composed of domains so that they can interact with multiple other molecules either simultaneously or sequentially 15:9 15.1 | The Basic Elements of Cell Signaling Systems Signals can combine in different ways to generate different outcomes A typical cell is exposed to hundreds of different signals In general, combinations of signals generate the different responses of cells 15:7  15.2 | A Survey of Extracellular Messengers and Their Receptors function as ligand- regulated transcription conduct a flow contain seven factors of ions across transmembrane α dimerize and activate their the plasma helices and cytoplasmic protein-kinase membrane to activate GTP- domain to phosphorylate change its binding proteins specific tyrosine residues of potential cytoplasmic substrate proteins 15:14 15.3 | Signal Transduction by G Protein-Coupled Receptors The G protein couples to a receptor: G protein-coupled receptor Different names – same protein complex G protein Heterotrimeric G protein Large G protein The G protein has 3 subunits: alpha (a), beta (b) , gamma (g) The alpha subunit is a GTPase: Homologous to Ran and Rho (GTPases discussed in nuclear transport and cytoskeletons) and all the other GTPases 15:15 15.3 | Signal Transduction by G Protein-Coupled Receptors Found in all eukaryotes, with over 700 GPCRs in humans Signaling molecules (ligands) vary: (proteins, peptides, amino acid derivatives, fatty acids, photons, olfactory molecules) Ligands activate receptors that stimulate effectors to give rise to a physiological response G protein-coupled receptors normally have their amino-terminus present on the outside of the cell, the seven a helices that traverse the plasma membrane connected by loops of varying length, and the carboxyl-terminus present on the inside of the cell 15:16  15.3 | Signal Transduction by G Protein-Coupled Receptors Receptors General structure of a G protein-coupled receptor 7 transmembrane α helices Ligand binding site Cytosolic portion that interacts with (large heterotrimeric) G proteins GRK phosphorylation sites for receptor downregulation PKA is activated by GPCR and also can participate in downregulation 15:18 15.3 | Signal Transduction by G Protein-Coupled Receptors Receptors: activation Ligand binding to the receptor extracellular domain changes the conformation of its intracellular domain What’s the GEF?(the guanine The receptor’s affinity for G proteins increases, and the nucleotide receptor binds the trimeric G protein exchange factor?) A GDP is exchanged for GTP on the Ga subunit which It’s the GPCR itself activates it and promotes association with the effector One ligand-bound receptor can activate many G proteins 15.3 | Signal Transduction by G Protein-Coupled Receptors Receptors: inactivation Termination of the response can occur through multiple processes Desensitization – by blocking active receptors from turning on additional G proteins (even though ligand is still bound) G protein-coupled receptor kinase (GRK) phosphorylates a GPCR Proteins called arrestins compete with G proteins to bind GPCRs If receptors are recycled and returned to the cell surface, the cells remain sensitive to the 15:20 ligand and are said to be resensitized  15.3 | Signal Transduction by G Protein-Coupled Receptors G proteins Hydrolysis of GTP to Ligand binding GDP in Ga results in induces dissociation with conformational effector and change in reassociation with Gbg receptor and binding of G protein to the receptor Ga binds to an effector protein Activated activating the receptor effector results in conformational change in Ga triggering Dissociation of dissociation of a subunit GDP Binding of GTP 15:21 15.3 | Signal Transduction by G Protein-Coupled Receptors What are the effector proteins? Membrane ion channels Enzymes that catalyze the formation of second messengers 21 different Gα subunits (encoded by 16 genes) 6 different Gβ subunits 12 different Gγ subunits 15:22 15.3 | Signal Transduction by G Protein-Coupled Receptors G-proteins can activate adenylyl cyclase Adenylyl cyclase removes two phosphates as pyrophosphate Reaction is driven in the forward direction by the hydrolysis of pyrophosphate cAMP is short-lived (unstable) Hydrolyzed to 5’-AMP 15:23  15.3 | Second Messengers The Discovery of Cyclic AMP Sutherland (Case Western Reserve University) developed an in vitro system to activate glycogen phosphorylase in cell extracts incubated with glucagon or epinephrine. Using centrifugation, glycogen phosphorylase was present only in the supernatant fraction, but the particulate material was required to obtain the hormone response. If the particulate fraction of a liver homogenate was isolated and incubated with the hormone, some substance was released that, when added to the supernatant fraction, activated the soluble glycogen phosphorylase molecules. Sutherland identified the substance released by membranes of the particulate fraction as cyclic adenosine monophosphate (cyclic AMP, or cAMP). This discovery is heralded as the beginning of the study of signal transduction. 15:24 First messenger = ligand Second messenger = small molecules that increase or decrease in concentration in response to first messenger (can be anywhere along the cascade) Second messengers bind to other proteins to modify their activity 15:25 Image illustrates some of the cell processes that can be affected by changes in cAMP concentration and activation of PKA by cAMP cAMP activates protein kinase A Protein kinase A (PKA) has many substrates Phosphorylates target proteins on: serine threonine Due to a variety of cell responses, PKA must phosphorylate different substrates, and over 100 substrates are known PKA phosphorylates the appropriate substrates in response to a particular stimulus, in a particular cell type 15:26 How can so many types of signals be mediated by PKA? One way is by confinement of the signaling process to one part of the cell via adaptor proteins A kinase anchoring protein (AKAP) AKAPs can be cell/tissue specific and confine PKA to actin filaments microtubules ion channels mitochondria nucleus AKAP5 functions as a scaffold protein (found in lipid rafts) - ”signaling hubs” note how it interacts with the regulatory subunits of PKA the end result is confinement of the PKA signal 15:27  AKAPS At least 50 different AKAPs have been discovered: provide a structural framework or scaffold for coordinating protein– protein interactions by sequestering PKA to specific cellular locations. Relevant substrates are present close by and are the first ones to become phosphorylated (i.e. substrate selection), and different cells express different AKAPs, resulting in localization of PKA in the presence of different substrates 15:28 Examples of PKA signaling pathways  15.3 | The Human Perspective Disorders Associated with G Protein-Coupled Receptors Over one‐third of all prescription drugs act as ligands that bind to this huge superfamily of receptors Several disorders are caused by defects in receptors or G proteins Loss of function mutations result in nonfunctional signal pathways Retinitis pigmentosa, a 2D representation of a “composite” progressive degeneration of the transmembrane receptor showing retina, can be caused my the approximate sites of a number mutations in rhodopsin’s ability to of mutations responsible for activate a G protein causing human diseases 15:29  15.3 | The Human Perspective Disorders Associated with G Protein- Coupled Receptors Gain of function mutations may create a constitutively activated G protein. Some benign thyroid tumors are caused by a mutation in a receptor. Certain polymorphisms in G protein- related genes may result in an increased susceptibility to asthma or high blood pressure, as well as decreased susceptibility to HIV. 15:29 15:30 Learning objectives: Know the 8 basic elements of cell signaling Know the 4 categories of intracellular signaling signals Understand the terms first messenger and second messenger Understand the principle of protein domains in docking Understand the principles of regulation by phosphorylation and dephosphorylation Understand the principles of GTPases Understand the basic principles of G proteins Understand the basic principles of G protein coupled receptors Understand the basic principles of signal transduction by GPCR Understand the role of GRK and arrestin Know the role and function of adenylyl cyclase and how cAMP is generated Be clear on the relationship between cAMP and PKA Understand the role of AKAP 15:31