i l principles and mechanisms€¦ · cell signaling: principles and mechanisms /wendell lim, bruce...
TRANSCRIPT
i I l I l t
I ,
I
principles and mechanisms
Wendell Lim
j Bruce Mayer
1 Tony Pawson l 1 l
. I
I l 1 I •
I , f"C\ Garland Science ~ Taylor&FrancisGroup
Garland Science Vice President: Denise Schanck Senior Editor: Michael Morales Production Editor: Natasha Wolfe Development Editor: Mary Purton Editorial Assistants: Lamia Harik and Alina Yurova Copy Editor: Jo Clayton Proofreader: Sally Huish 1 Illustrations and Design: Matthew Me · Indexer: Medical Indexing Ltd. Typesetter: Thomson Digital
F.~ LIM D'H725
© 2015 by Garland Science, Taylor & Francis Group, LLC
Front Cover: Cover painting and chapter-opening images by the British artist and architect Victor Pasmore (1908-1998). Used with permission of The Victor Pasmore Estate. Copyright is with the Estate of the Artist.
I.
This book contains information obtained from authentic and highly regarded sources. Every effiJI't has been made to trace copyr.igh holders and to obtah1 their permission for the use of copyl'ight materia l. Reprinted material is quoted with permission, and sources are indicated. A wide variety of references are listed. Reasonable efforts have been made to publish reliable data and information, but the author and the publisher cannot assume responsibility for the validity of all materials or for the consequences of their use.
All rights reserved. No part of this book covered by the copyright hereon may be reproduced or used in any format in any form or by any means- graphic, electronic or mechanical, including photocopying, recording, taping, or informat ion storage and retrieval systems-without permission of the publisher.
ISBN 978-0-8153-4244-1
Published by Garland Science, Taylor & Francis Group, LLC, an informa business, 711 Third Avenue, New York, NY, 10017, USA, and 3 Park Square, Milton Park, Abingdon, OX14 4RN, UK.
Printed in the United States of America
15 14 13 12 1110 9 8 7 6 54 3 2 1
GS Garland Science Taylor & Francis Group
Visit our Web site at http://www.garlandscience.com
Wendell Lim is at the University of California, San Francisco, where he is Professor of Cellular and Molecular Pharmacology. The principal focus of his resear ch is the s tructure and mechanism of protein interaction domains and the logic by which these components are LlSed to build complex cellular signaling systems.
Bruce Mayer is at the University of Connecticut Health Center, where he is a P•·ofessor of Genetics and Developmental Biology. Current work in his group focuses on characterizing and manipulating tyrosine kinase-mediated signal transduction pathways.
Tony Pawson, deceased, was Senior Investigator and Director of Research in the Department of Medical Genetics and Microbiology at the University of Toronto where his research focused on the molecular dissection and functional significance of proteinprotein interactions in signal transduction.
Library of Congress Cataloging-in-Publication Data
Lim, Wendell, author. Cell signaling: principles and mechanisms /Wendell Lim, Bruce
Mayer, Tony Pawson. pages em
ISBN 978-0-8153-4244-1 (paperback) 1. Cell interaction. 2. Cellular signal transduction. I. Mayer,
Bruce, author. II. Pawson, T., author. III. Title. QH604.2.L56 2014 571.7'4--dc23
2014010851
This ~oak is de~icated to the memory of Tony Pawson (1952-2013). Tony was a towerzng figure zn the field of cell signaling, and he will be sorely missed. The photo was taken at the Gerstle Park Inn in San Rafael, CA, where we often met to work on the book under the spreading branches of this ancient live oak tree.
Brief Contents
Chapter 1: Introduction to Cell Signaling
Chapter 2: Principles and Mechanisms of Protein Interactions
Chapter 3: Signaling Enzymes and Their Allosteric Regulation
Chapter 4: Role of Post-Translational Modifications in Signaling
Chapter 5: Subcellular Localization of Signaling Molecules
Chapter 6: Second Messengers: Small Signaling Mediators
Chapter 7: Membranes, Lipids, and Enzymes That Modify Them
Chapter 8: Information Transfer Across the Membrane
Chapter 9: Regulated Protein Degradation
Chapter 1 0: The Modular Architecture and Evolution of Signaling Proteins
Chapter 11: Information Processing by Signaling Devices and Networks
Chapter 12: How Cells Make Decisions
Chapter 13: Methods for Studying Signaling Proteins and Networks
Glossary
Index
1
21
43
85
115
135
155
177
217
243
275
305
345
375
385
Detailed Contents
Chapter 1 Introduction to Cell Signaling 1
WHAT IS CELL SIGNALING? 1
All cells have the ability to respond to their environment 2
Cells must perceive and respond to a wide range of signals 3 Signaling systems need to solve a number of
common problems 4
THE FUNDAMENTAL ROLE OF SIGNALING IN BIOLOGICAL PROCESSES 6
Work in many different fields converged to reveal the underlying mechanisms of signaling 6
Despite the diversity of signaling pathways and mechanisms, fundamental commonalities have emerged 7
Signaling must operate at multiple scales in space and time 9
THE MOLECULAR CURRENCIES OF INFORMATION PROCESSING
Information is transferred by changes in the state of proteins
There is a limited number of ways in which the state of proteins can change
Most changes in state involve simultaneous changes in several different currencies
LINKING SIGNALING NODES INTO PATHWAYS AND NETWORKS
Information transfer involves linking different changes of state together
Multiple state changes are linked together to generate pathways and networks
Cellular information-processing systems have a hierarchical architecture
Summary
Questions
References
Chapter 2
11
11
12
14
15
15
16
17
18
18
19
The affinity and specificity of an interaction determine how likely it is to occur in the cell
The strength of a binding interaction is defined by the dissociation constant (I\u)
The dissociation constant is related to the binding energy of the interaction
The dissociation constant is also related to rates of binding and dissociation
PROTEIN INTERACTIONS IN THEIR CELLULAR AND MOLECULAR CONTEXT
The apparent dissociation constant can be strongly affected by the local cellular environment and other binding partners
Ideal affinity and specificity depends on biological function and ligand concentrations
There are functional constraints on interaction affinities and specificities
Interaction affinity and specificity can be independently modulated
Cooperativity involves the coupled binding of multiple ligands
Diverse molecular mechanisms underlie cooperativity
Cooperative binding has a variety of functional consequences
Protein assemblies differ in their stability and homogeneity
Summary
Questions
References
Chapter 3 Signaling Enzymes and Their Allosteric Regulation
PRINCIPLES OF ENZYME CATALYSIS
Enzymes have a number of properties that make Principles and Mechanisms of Protein Interactions
PROPERTIES OF PROTEIN-PROTEIN INTERACTIONS
them useful for transmitting signals in the cell 21 Enzymes use a variety of mechanisms to
Changes in protein binding have both direct and indirect functiona l conseqt1ences
Pr~tein binding can b mediated by broad mteraction surfaces or by short, linear peptides
enhance the rate of chemical reactions
22 Enzymes can drive reactions in one direction by energetic coupling
22 ALLOSTERIC CONFORMATIONAL CHANGES
Conformational flexibility of proteins enables 23 allosteric control
24
25
27
28
30
30
31
32
34
35
36
36
37
38 38 40
43
44
44
45
46
47
47
I I
Detailed Contents Detailed Contents
Signaling proteins employ diverse classes of Questions 80 Chapter 5 Small signaling mediators are controlled by an
conformational rearrangements 48 References 82 Subcellular Localization of Signaling interplay of their production and elimination 136
PROTEIN PHOSPHORYLATION AS Molecules 115 Small signaling mediators exert their effects
A REGULATORY MECHANISM 49 Chapter4 LOCALIZATION AS A SIGNALING CURRENCY 115 by binding downstream effectors 136
Phosphorylation can act as a regulatory mark 49 Role of Post-Translational Changes in subcellular localization can Small signaling mediators can lead to fast, Modifications in Signaling 85 transmit information distant, and amplified signal transmission 137
Phosphorylation can either disrupt or 116
induce protein structure 50 THE LOGIC OF POST-TRANSLATIONAL Subcellular localization can be regulated Small signaling mediators can generate
PROTEIN KINASES 52 REGULATION 85 by a variety of mechanisms 117 complex temporal and spatial patterns 138
The structure and catalytic mechanism of protein Proteins can be covalently modified by CONTROL OF NUCLEAR LOCALIZATION 117 CLASSES OF SMALL SIGNALING MEDIATORS 139
kinases are conserved 52 the addition of simple functional groups 86 Short, modular peptide motifs direct nuclear Small signaling mediators have a wide range of
The activation loop and C-helix are conserved Proteins can also be covalently modified by the import and export 118 physical properties 140
molecular levers that conformationally addition of sugars, lipids, and even proteins 87 Nuclear transport is controlled by shuttle The cyclic nucleotides cAMP and cGMP are
control kinase activity 54 Post-translational modifications can alter proteins and the G protein Ran 118 produced by cyclase enzymes and destroyed
Insulin receptor kinase activity is controlled protein structure, localization, and stability 88 Phosphorylation of transcription factor Pho4 by phosphodiesterases 140
via activation-loop phosphorylation 54 Post-translational control machinery often regulates nuclear import and export 119 Cyclic nucleotides regulate diverse cellular activities 141
Phosphorylation mediates long-range works as part of"writer/eraser/reader" systems 90 Nuclear import of STATs is regulated by The regulatory (R) subunit of protein kinase
conformational regulation of Src family kinases 55 Post-translational modifications allow very rapid phosphorylation and conformational change 120 A is a conformational sensor of cAMP binding 142
Multiple binding interactions regulate signaling and transmission of spatial information 92 Localization of MAP kinases is regulated Some small signaling mediators are
protein kinase substrate specificity 56 INTERPLAY BETWEEN POST-TRANSLATIONAL by association with nuclear and cytosolic derived from membrane lipids 143
Protein kinases can be divided into nine families 58 MODIFICATIONS 92 binding partners 121 PLC generates two signaling mediators,
PROTEIN PHOSPHATASES 60 A post-translational modification can Notch nuclear localization is regulated by IP3 and DAG 144
Serine/threonine phosphatases are metalloenzymes 60 promote or antagonize other modifications 93 proteolytic cleavage 122 Activation of protein kinase C is regulated
Most tyrosine phosphatases utilize a catalytic p53 is tightly regulated by a wide variety CONTROL OF MEMBRANE LOCALIZATION 122
by IP3 and DAG 144
cysteine residue 62 of post-translational modifications 95 Proteins can span the membrane or be CALCIUM SIGNALING 145
Tyrosine phosphatases are regulated by modular The level and activity of p53 are regulated associated with it peripherally 122 Activation of Ca2• channels is a common
domains while serine/threonine phosphatases by ubiquitylation and acetylation 96 Proteins can be covalently modified with means ofregulation 146
often associate with regulatory accessory subunits 64 Additional modifications further fine-tune p53 activity 96 lipids after translation 123 Ca2+ influx is rapid and local 147
G PROTEIN SIGNALING 65 PROTEIN PHOSPHORYLATION 97 Modular lipid-binding domains are important for Calmodulin is a conformational sensor of
G proteins are conformational switches Phosphorylation is often coupled with protein regulated association of proteins with membranes 124 intracellular calcium levels 147
controlled by two opposing enzymes 65 interactions 97 Some lipid-modified proteins can reversibly Signaling can lead to propagating Ca2• waves 148
The presence of the GTP y-phosphate determines Kinases and phosphatases vary in their associate with membranes 125 SPECIFICITY AND REGULATION
Coupling effector protein activation to 149
the structure of G protein switch I and II regions 66 substrate specificity 99 Scaffold proteins can increase input and output membrane recruitment is a common theme
There are two major classes of signaling G proteins 67 Multiple phosphorylation of proteins can in signaling
specificity of small-molecule signaling 150
Subfamilies of small G proteins regulate arise by different mechanisms 100 126 AKAP scaffold proteins can also regulate dynamics
Akt kinase is regulated by membrane recruitment diverse biological functions 67 Histidine and other amino acids can be
and phosphorylation of cAMP signaling 150
Many upstream receptors feed into a small set of phosphorylated, especially in prokaryotes 101 126 Summary
MODULATION OF SIGNALING BY 152
common heterotrimeric G proteins 68 Two-component systems and histidine Questions phosphorylation are also present in eukaryotes 103 MEMBRANE TRAFFICKING 127
152
REGULATORY ENZYMES FOR G PROTEIN Proteins can be internalized by a variety References 153
SIGNALING 70 ADDITION OF UBIQUITIN AND RELATED PROTEINS 104
of mechanisms 127 Chapter 7 G-protein-coupled receptors act as GEFs Internalization of receptors can modulate
for heterotrimeric G proteins 71 Specialized enzymes mediate the addition signal transduction Membranes, Lipids, and Enzymes
and removal of ubiquitin 104 128 That Modify Them 155 Distinct GEF and GAP domains regulate TGFp signaling output depends on the specific small G protein families 71 E3 ubiquitin ligases determine which proteins mechanism of receptor internalization BIOLOGICAL MEMBRANES AND THEIR
GEFs catalyze GDP/GTP exchange by will be ubiquitylated 105 129 PROPERTIES
Retrograde signaling allows effects distant 155 deforming the nucleotide-binding pocket 73 Ubiquitin-binding domains read ubiquitin-mediated from the site of ligand binding Biological membranes consist of a variety of polar lipids 156
signals in diverse cellular activities 106 130 GAPs order the catalytic machinery for hydrolysis 74 Ras isoforms in distinct subcellular locations Structural properties of membrane lipids favor
Regulators of G protein signaling (RGS) proteins HISTONE ACETYLATION AND METHYLATION 107 have different signaling outputs 130 the formation ofbilayers 157
act as GAPs for heterotrimeric G proteins 75 Chromatin structure is regulated by post- Summary The composition of the membrane determines 132
Additional mechanisms are used to translational modification ofhistones Questions its physical properties 158
fine-tune the activity of G proteins 75 and associated proteins 108 132 There are fundamental differences between References
SIGNALING ENZYME CASCADES 75 Two writer/eraser/reader systems are based on 132 biochemistry in solution and on the membrane 160
The three-tiered MAP kinase cascade forms protein methylation and acetylation 109 Chapter 6 LIPID-MODIFYING ENZYMES USED
a signaling module in all eukaryotes 76 Chromatin modification in transcription is dynamic Second Messengers: Small Signaling
IN SIGNALING 161
Scaffold proteins often organize MAPK cascades 77 and leads to highly cooperative interactions 110 Mediators Cleavage of membrane lipids by phospholipases Summary 135
G protein activity can also be regulated by 112 PROPERTIES OF SMALL SIGNALING
generates a variety of bioactive products 161
signaling cascades 79 Questions 113 A variety oflipid kinases and phosphatases
Summary 80 References 114 MEDIATORS 135 are involved in signaling 163
• Detailed Contents Detailed Contents
EXAMPLES OF MAJOR LIPID SIGNALING The voltage-gated potassium channel provides clues Questions 241 Certain plant protein kinases are regulated by
PATHWAYS 164 to mechanisms of gating and ion specificity 200 References 242 modular light-gated domains 267
Phosphoinositides can serve as membrane binding Ligand-gated ion channels play a central Regulation of the neutrophil NADPH oxidase
sites and as a source of signaling mediators 164 role in neurotransmission 202 Chapter 10 by modular interactions 268
Phosphoinositide species provide a set of membrane MEMBRANE-PERMEABLE SIGNALING 204 The Modular Architecture and CREATING NEW FUNCTIONS THROUGH
binding signals 166 Nitric oxide mediates short-range signaling Evolution of Signaling Proteins 243 DOMAIN RECOMBINATION 269
Phospholipase D generates the important in the vascular system 204 MODULAR PROTEIN DOMAINS 244 Some modular domain rearrangements can lead
signaling mediator, phosphatidic acid (PA) 168 02
binding regulates the response to hypoxia 205 Protein domains usually have a globular structure 244 to cancer 269
Phospholipase D plays a role in mTOR signaling 169 The receptors for steroid hormones are Bioinformatic approaches can identifY protein domains 244 Modules can be recombined experimentally
The metabolism of sphingomyelin generates transcription factors 206 Domains can be composed of several smaller repeats 245 to engineer new signaling behaviors 270
a host of signaling mediators 170 DOWN-REGULATION OF RECEPTOR SIGNALING 208 Protein domains often act as recognition modules 246 Summary 272
Phospholipase A2 generates the precursor for Ubiquitylation regulates the endocytosis, recycling, INTERACTION DOMAINS THAT RECOGNIZE Questions 272 a family of potent inflammatory mediators 172
and degradation of cell-surface receptors 209 POST-TRANSLATIONAL MODIFICATIONS 249 References 273 Summary 174 G protein coupled receptors are desensitized SH2 domains bind phosphotyrosine-containing sites 249 Questions 174 by phosphorylation and adaptor binding 211 Some SH2 domains are elements oflarger binding
Chapter 11
References 174 Summary 213 structures 252 Information Processing by Signaling Devices and Networks 275
Questions 213 Several different types of interaction domains Chapter 8
References 215 recognize phosphotyrosine 252 SIGNALING SYSTEMS AS INFORMATION-Information Transfer Across the Membrane 177 Multiple domains recognize motifs phosphorylated PROCESSING DEVICES 276
PRINCIPLES OF TRANSMEMBRANE SIGNALING 177 Chapter 9 on serine/threonine 254 Signaling devices can be considered as state machines 276
The cell must process and respond to a Regulated Protein Degradation 217 14-3-3 proteins recognize specific phosphoserine/ Signaling devices are organized in a hierarchical
diversity of environmental cues 178 GENERAL PROPERTIES AND EXAMPLES OF phosphothreonine motifs 254 fashion 277
Three general strategies are used to SIGNAL-REGULATED PROTEOLYSIS 217 Interaction domains recognize acetylated and Signaling devices face a variety of challenges in
transfer information across the membrane 179 Proteases are a diverse group of enzymes 218 methylated sites 255 input detection 278
Many drugs target receptors 180 Blood coagulation is regulated by a Ubiquitylation regulates protein-protein interactions 256 Proteins can function as simple signaling devices 279
TRANSDUCTION STRATEGIES USED BY cascade of proteases 219 INTERACTION DOMAINS THAT RECOGNIZE INTEGRATING MULTIPLE SIGNALING INPUTS 281
TRANSMEMBRANE RECEPTORS 180 Regulated proteolysis by metalloproteases UNMODIFIED PEPTIDE MOTIFS OR PROTEINS 257 Logic gates process information from multiple inputs 281
Receptors with multiple membrane-spanning can generate signaling molecules and Proline-rich sequences are favorable Simple peptide motifs can integrate multiple
segments undergo conformational changes alter the extracellular environment 220 recognition motifs 257 post-translational modification inputs 282
upon ligand binding 180 ADAMs regulate signaling pathways SH3 domains bind proline-rich motifs 258 Cyclin-dependent kinase is an allosteric
Receptors with a single membrane-spanning segment by cleaving membrane-associated proteins 221 PDZ domains recognize C-terminal peptide motifs 258 signal-integrating device 283
form higher-order assemblies upon ligand binding 181 MMPs participate in remodeling the Protein interaction domains can form dimers Modular signaling proteins can integrate multiple
Receptor clustering confers advantages for extracellular environment 222 or oligomers 259 inputs 284
signal propagation 182 Proteolysis activates the thrombin receptor 223 Transcriptional promoters can integrate input from INTERACTION DOMAINS THAT RECOGNIZE
G-PROTEIN-COUPLED RECEPTORS 184 Regulated intramembrane proteolysis (RIP) is PHOSPHOLIPIDS 260 multiple signaling pathways 285
G-protein-coupled receptors have intrinsic an essential step in signaling by some receptors 224 RESPONDING TO THE STRENGTH OR PH domains form a major class of phosphoinositide-
enzymatic activity 184 UBIQUITIN AND THE PROTEASOME binding domains 260 DURATION OF AN INPUT 286
Signaling by GPCRs can be very fast and DEGRADATION PATHWAY 225 FYVE domains are phospholipid-binding domains Signaling systems can respond to signal amplitude
lead to enormous signal amplification 186 The proteasome is a specialized molecular machine found in endocytic proteins 261 in a graded or a digital manner 288
TRANSMEMBRANE RECEPTORS ASSOCIATED that degrades intracellular proteins 225 BAR domains bind and stabilize curved membranes 262 An enzyme can behave as a switch through
WITH ENZYMATIC ACTIVITY 186 The cell cycle is controlled by two large CREATING COMPLEX FUNCTIONS BY
cooperativity 289
ubiquitin-conjugating complexes 226 Networks can also yield switchlike activation 290 Receptor tyrosine kinases control important cell fate COMBINING INTERACTION DOMAINS 262
decisions in multicellular eukaryotes 186 SCF recognizes specific phosphorylated proteins, Recombination of domains occurs through evolution 262
Signaling systems can distinguish between targeting them for destruction 227 transient and sustained input 292
TGF~ receptors are serine/threonine Combinations of interaction domains or motifs kinases that activate transcription factors 187 Two APC species act at distinct points in
can be used as a scaffold for the assembly of MODIFYING THE STRENGTH OR DURATION OF the cell cycle 228 OUTPUT Some receptors have intrinsic protein signaling complexes 263 294
phosphatase or guanylyl cyclase activity 189 NF-KB is controlled by regulated Scaffold proteins containing PDZ domains organize Signaling pathways often amplify signals as they
Noncovalent coupling of receptors to protein degradation of its inhibitor 230 cell-cell signaling complexes such as the are transmitted 294
kinases is a common signaling strategy 189 CASPASE-MEDIATED CELL DEATH PATHWAYS 232 postsynaptic density 264 Negative feedback allows fine-tuning of output 294
Some receptors use complex activation Apoptosis is an orderly and highly regulated form Proteins with multiple phosphotyrosine motifs Adaptation allows cells to control output duration 296 pathways that involve both kinase activation of cell death 232 function as dynamically regulated scaffolds 265 Feedback can cause output levels to oscillate and proteolytic processing 193 The activity of caspases is tightly regulated 233
RECOMBINING INTERACTION AND CATALYTIC between two stable states 299 Wnt and Hedgehog are two important The extrinsic pathway links cell death receptors DOMAINS TO BUILD COMPLEX ALLOSTERIC Bistable responses also underlie more permanent
signaling pathways in development 194 to caspase activation 235 SWITCH PROTEINS outputs 301 266 A variety of receptors couple to proteolytic activities 197 Mitochondria orchestrate the intrinsic cell death Many signaling enzymes are allosteric switches Summary 303 266 GATED CHANNELS 199 pathway 238 14-3-3 Protein regulates the Rafkinase by Questions 303 Gated channels share a similar overall structure 199 Summary 241 coordinately binding two phosphorylation sites 267 References 303
Detailed Contents
Chapter 12 How Cells Make Decisions 305
VERTEBRATE VISION 307
Organ: vertebrate eye 308
Cell: photoreceptor cell 308
Molecular Network: visual transduction cascade 309
How does the photoreceptor cell detect light and convert it to a biochemical signal? 310
How is the photoreceptor cell able to detect low light, even a single photon? 311
How can the response be so rapid? 312
How does the photoreceptor cell reset itself to enable detection of further changes in light? 312
Summary 314
References 314
PDGF SIGNALING 315
Tissue: process of wound healing 316
Cell: fibroblast response to wounding & platelet activation 316
Molecular Network: control of fibroblast proliferation 317
How do fibroblasts detect the local occurrence of a wound? 318
How are PDGF signals propagated within the cell to generate outputs such as cell proliferation? 319
How is misactivation of the proliferation response prevented? 320
How is the proliferative response terminated? 321
Summary 322
References 322
THE CELL CYCLE 323
Cell: distinct phases of the cell cycle 324
Molecular Network: cyclin-dependent kinase (Cdk) is a central switch whose activity is modulated by the different cyclins 325
How are sharp and commited transitions between cell cycle phases achieved? 326
How does the cell cycle ensure that each transition proceeds only under appropriate conditions? 329
Summary 331
References 331
T CELL SIGNALING 333
Organism: launching the adaptive immune response 334
Cell: engagement ofT cell and antigen presenting cell 335
Molecular Network: T cell receptor (TCR) signaling network 336
How does the T cell receptor transmit signals after peptide/MHC recognition? 338
How does the T cell prevent misactivation? 339
How does the T cell launch a robust response when stimulated by as few as ten antigenic peptide complexes? 340
How might the T cell discriminate between antigenic and non-antigenic peptides? 342
Summary 344
References 344
Chapter 13 Methods for Studying Signaling Proteins and Networks
BIOCHEMICAL AND BIOPHYSICAL ANALYSIS OF PROTEINS Analytical methods can determine quantitative
binding parameters Michaelis-Menten analysis provides a way to
measure the catalytic power of enzymes Methods to determine and analyze protein
conformation are central to the study of signaling
X-ray crystallography provides high-resolution protein structures
Nuclear magnetic resonance (NMR) can reveal the dynamic structure of small proteins
Electron microscopy can map the shape of very large protein complexes
Specialized spectroscopic methods can be used to study protein dynamics
MAPPING PROTEIN INTERACTIONS AND LOCALIZATION Interacting proteins can be identified by
isolating protein complexes from cell extracts Binding partners can be identified by screening
large libraries of genes Direct protein-protein interactions can be detected
by solid-phase screening Fluorescent protein tags are used to locate and
track proteins in living cells Protein-protein interactions can be visualized
directly in living cells
METHODS TO PERTURB CELL SIGNALING NETWORKS AND MONITOR CELLULAR RESPONSES Genetic and pharmacological methods can be used
to perturb networks Chemical dimerizers and optogenetic proteins provide a
dynamic way to artificially activate pathways eDNA microarrays and high-throughput
sequencing are used to monitor the transcriptional state of a cell
Modification-specific antibodies provide a method to track post-translational changes
Mass spectrometry is the workhorse for identification of proteins and their modifications
Live-cell time-lapse microscopy provides a way to track the dynamics of single-cell responses
Biosensors allow signaling activity to be monitored in living cells
Flow cytometry provides a method to analyze rapidly single-cell responses
Questions
References
Glossary
Index
345
345
345
347
349
352
353
353
354
355
355
356
357
357
359
360
361
362
363
364
366
368
369
371
372
373
375
385
•'
Introduction to Cell Signaling
All living cells perceive signals from the outside environment and adjust their behavior accordingly. If you think back to the earliest living cells, it is easy to imagine the incredible pressure they were under to evolve the ability to sense features of the environment and to change in response to these signals. The ability to sense and move toward nutrients, or to sense and avoid stresses and toxins, would give a unicellular organism a huge competitive advantage. This ability to respond to environmental cues is important for single cells, but it is also absolutely essential for the normal development and functioning of multicellular organisms, which depend on a continuous and extensive exchange of information to coordinate the activities of many individual cells. Furthermore, when this cellular communication goes awry, it can result in diseases such as cancer. In this chapter, we will introduce the basic principles of cell signaling and the molecular mechanisms that underlie it.
WHAT IS CELL SIGNALING?
Cells are the smallest fundamental units of life. Part of what makes them so distinctly "living" is their remarkable ability to sense stimuli and to respond to them in a dynamic manner. This ability of cells to detect or receive information and process it to make decisions can also be considered from the broader perspective of information processing. Here we can draw analogies to the engineering and design principles of other, more familiar information-processing systems, such as humanmade electronic devices. It is this interface between the unique properties of living systems and the more universal properties of any system that processes information that makes the study of cellular signaling mechanisms so compelling.