_Current Topics in Membranes, Volume 4 9

Gap junctionsMolecular Basis of Cel l

Communication in Health

and Disease

Camillo Peracchia

Contributors xviiPreface xxiiiPrevious Volumes in Series xx v

PART 1 Channel Structure, Assembly, and Degradation

CHAPTER 1 Gap Junction Structure : New Structures andNew InsightsGina Sosinsky

I. Overview of Gap Junction Structure 2II. The Constituent Proteins of Gap Junctions :

Size and Topology Models of the Connexi nFamily 4

III. Isolation and Purification of Gap Junctions 6

IV. Molecular Structure of Gap Junctions Determined byX-Ray Diffraction and Electron Microscopy 7

V. Concluding Remarks 1 7References 18

CHAPTER 2 Degradation of Gap Junctions and ConnexinsJames G. Laing and Eric C. Beyer

I. Most Connexins Turn Over Rapidly 2 4II. Ubiquitin Pathway and Pathways of Protei n

Degradation 25

III. Ubiquitin Dependence of Cx43 Degradation 2 9IV. Membrane Protein Degradation 30V. Lysosomal and Proteasomal Degradation o f

Cx43 31VI. Phosphorylation and Regulation of Connexin

Degradation 34

VII. Heat-Induced Degradation of Cx43 3 5

VIII. Conclusion 36References 37

PART II Channel Forms, Permeability, and Conductance

CHAPTER 3 Homotypic, Heterotypic, and Heteromeric GapJunction ChannelsP. R. Brink, V. Valiunas, and G . J. Christ

I. Introduction 43II. Homotypic hCx37 and rCx43 Gap Junction Channels

III. Heterotypic hCx37-rCx43 Gap Junction Channels 46IV. Co-transfection of hCx37 and rCx43 : Heteromeric Gaj

Junction Channels 4 9V. Why Would a Cell Bother with Heteromeric Ga p

Junction Channels? 57References 58

CHAPTER 4 Heteromultimeric Gap Junction Channels andCardiac Diseas eSergio Elenes and Alonso P. Moreno

I. Introduction 6 1II. Gap Junctions : Structure and Nomenclature 62

III. Endogenous Expression of Multiple Connexins i nVarious Tissues 64

IV. Experimental Formation of Heteromultimeric Channehin Exogenous Systems 67

V. Molecular Regions Involved in Assembly 8 0VI. Physiological Implications of Heteromultimeric Chann e

Formation 82VII. Conclusions and Future Directions 87

References 87

CHAPTER 5 Ion Permeation through Connexin Gap Junctio nChannels : Effects on Conductance and Selectivit yRichard D. Veenstra

I. Introduction 9 5II. Theories of Electrodiffusion 99

III. Gap Junction Channel Conductance an dPermeability 109

IV. Summary 124References 125

CHAPTER 6 Phosphorylation of Connexins : Consequences fo rPermeability, Conductance, and Kinetics of Ga pJunction ChannelsHabo J. Jongsma, Harold V. M. van Rijen, Brenda R. Kwak,and Marc Chanson

I . Introduction 13 1H. Connexin43 13 2

III. Connexin40 and -45 13 6IV. Connexin26 and -32 13 9

V. Concluding Remarks 14 0References 142

CHAPTER 7 Intercellular Calcium Wave Communication via GapJunction-Dependent and -Independent Mechanism sEliana Scemes, Sylvia O. Suadicani, and David C. Spray

L Introduction 145II. Two Routes for Intercellular Calcium Wave

Propagation 14 7III. Some Features of Intercellular Ca Z+ Waves Depend

upon the Initiating Stimulus 14 8IV. Mechanisms for Intercellular Ca t+ Wav e

Propagation 15 5V. How Connexins Can Potentially Influence and Modulat e

the Propagation of Intercellular CaZ ` Waves 15 9VI. How the Extracellular Space May Influence Calcium

Wave Propagation 161VII . Functional Roles of Intercellular Calcium Waves 16 2

VIII . Prospects 16 6References 166

PART III Voltage Grating

CHAPTER 8 Membrane Potential Dependence of Gap Junctionsin VertebratesLuis C. Barrio, Ana Revilla, Juan M . Gomez-Hernandez,Marta de Miguel, and Daniel Gonzalez

I. Membrane Potential Dependence Is a Commo nRegulatory Mechanism among the Gap Junctions ofVertebrates 175

II. One Mechanism of Vm Gating Resides in EachHemichannel 18 0

III. A Gating Model of Junctions with Combined Vi and V,,,Dependence 18 2

IV. Functional Role of Vm Dependence 18 5References 186

CHAPTER 9 A Reexamination of Calcium Effects on Gap Junction sin Heart MyocytesBruno Delage and Jean Deleze

I. Introduction 18 9II. The Calcium Hypothesis: Is Cell Coupling Regulated by

Ca' Ions? 191III. Cytosolic Calcium Levels Correlating with Electrica l

Uncoupling 192IV. Conclusions 202

References 20 3

PART IV Chemical Grating

CHAPTER 10 Distinct Behaviors of Chemical- and Voltage-Sensitive Gates of Gap Junction Channe lFeliksas F. Bukauskas and Camillo Peracchia

I. Introduction 207II. C0 2 -Induced Gating at Different Vi 's 208

III. Channel Reopening in Response to Reversal of ViPolarity 212

IV. Kinetics of Unitary Transitions 21 6V. Conclusions 21 8

References 21 9

CHAPTER 11 A Molecular Model for the Chemical Regulationof Connexin43 Channels: The "Ball-and-Chain"Hypothesi sMario Delmar, Kathleen Stergiopoulos, Nobuo Homma,Guillermo Calero, Gregory Morley, Jose F. Ek-Vitorin,and Steven M. Taffe t

I. Introduction 22 3II. Connexin, the Gap Junction Protein 224

III. pH Regulation of Connexins 225IV. Regulation of Cx43 by Protein Kinases 22 6V. Structure-Function Studies on pH Gating of

Cx43 22 7VI. The Particle-Receptor Concept Put in Practice : Peptide

Block of pH Gating of Cx43 23 1

VII. Applicability of the Particle-Receptor Model to Ga pJunction Regulation by Other Factors 23 4

VIII. Cx43 Concatenants Do Not Function as the SimpleAddition of Individual Subunits 237References 24 3

CHAPTER 12 Mechanistic Differences between Chemical an dElectrical Gating of Gap Junction sL M. Skerrett, J. F. Smith, and B. J . Nicholso n

L Introduction 249II. The Voltage Gating Mechanism 253

III. Chemical Gating 262IV. Conclusions 265

References 26 6

CHAPTER 13 Behavior of Chemical- and Slow Voltage-SensitiveGates of Connexin Channels : The "Cork"Gating Hypothesi sCamillo Peracchia, Xiao G . Wang, and Lillian L. Peracchia

I. Introduction 27 1II. Role of Cytosolic pH and Calcium in Channe l

Gating 272III. Potential Participation of Calmodulin in the Gatin g

Mechanism 274IV. Connexin Domains Relevant to pHlCa Gating 27 5V. Does Chemical Gating Require Connexi n

Cooperativity? 278VI. Is the Chemical Gate Voltage Sensitive? 27 9

VII. Are There Intramolecular Interactions Relevant t oGating? 287

VIII. The "Cork" Gating Model 288References 291

CHAPTER 14 Molecular Determinants of Voltage Gating of Ga pJunctions Formed by Connexin32 and 26Thaddeus A. Bargiello, Seunghoon Oh, Yi Ri,Priscilla E. Purnick, and Vytas K. Verselis

I. Introduction 29 7II. Vi -Dependent Gating 298

III. Molecular Determinants of Vj Gating 302IV. Structural Implications 305V. Role of P87 Vj Gating 306

VI. Conclusions 31 1References 31 2

CHAPTER 15 Regulation of Connexin43 by TyrosineProtein KinasesAlan F Lau, Bonnie Warn-Cramer, and Rui Lin

I . Introduction 31 5IL Regulation of Cx43 by Nonreceptor Tyrosin e

Kinases 31 6III. Regulation of Cx43 by Receptor Tyrosine Kinases 32 5IV. The "Particle-Receptor" Model of Phosphorylation -

Induced Cx43 Channel Closure 333V. Summary and Future Directions 334

References 33 7

CHAPTER 16 Gating of Gap Junction Channels and Hemichannel sin the Lens : A Role in Cataract?Reiner Eckert, Paul Donaldson, JunSheng Lin, Jacqui Bond ,Colin Green, Rachelle Merriman-Smith, Mark Tunstall, an dJoerg Kistle r

I. Introduction 34 3II. The Lens Circulation System and Role of Gap Junctio n

Channels 344

III. Molecular Composition and Functional Properties ofLens Gap Junction Channels 34 6

IV. pH-Sensitive Gating of Lens Fiber Gap Junctions 34 7V. Fiber Cell Currents Reminiscent of Gap Junctio n

Hemichannels 35 1VI. A Role for Gap Junction Channels and Hemichannels i n

Cataract? 352References 353

PART V Hemichannels

CHAPTER 17 Biophysical Properties of Hemi-Gap-Junctiona lChannels Expressed in Xenopus OocytesL. Ebihara and J. Pal

I. Introduction 35 7II. Expression of Rat Cx46 in Xenopus Oocytes 359

III. Single Channel Properties of Cx46 Hemichannels 36 1IV. Voltage Gating for Cx46 Hemichannels and Cx4 6

Hemichannels in Intercellular Channels 36 1V. Structure of Pore Lining Region of Cx46 Hemichannel s

Inferred from Cysteine Scanning Mutagenesis 36 2VI. Properties of Hemichannels Formed from Different

Connexins 36 3VII. Heteromeric Association of Connexins Modifie s

Hemichannel Behavior 363VIII. Summary and Conclusions 364

References 36 5

CHAPTER 18 Properties of Connexin50 Hemichannels Expressed inXenopus laevis OocytesSepehr Eskandari and Guido A . Zampighi

I. Introduction 369II. Experimental Procedures 37 1

III. Electrophysiological Studies of Oocytes ExpressingConnexin50 373

IV. Morphological Studies of Oocytes Expressin gConnexin50 37 8

V. Conclusions 386References 38 6

PART VI Invertebrate Gap Junction s

CHAPTER 19 Gap Junction Communication in Invertebrates :The Innexin Gene FamilyPauline Phelan

I. Introductory Note 390II. Searching for Gap Junction Genes and Proteins in

Invertebrates 390

HI . Innexins : Functional Connexin Analogues in Drosophilaand C. elegans 393

IV. Genetic Screens Unwittingly Identified Gap Junctio nMutants 39 3

V. Cloning Defined a New Gene Family with No Homologyto the Vertebrate Connexins 402

VI. Innexin Proteins 405VII. Functional Expression of Innexins in Heterologous

Systems 408VIII. Distribution of Innexins 41 1

IX. Innexins and the Study of Gap Junction Function i nInvertebrates 412

X. Looking Forward 415References 41 6

PART VII Diseases Based on Defects of

Cell Communicatio n

CHAPTER 20 Hereditary Human Diseases Caused by

Connexin Mutation sMichael V. L . Bennett and Charles K. Abram s

I. Introduction 42 3

II. Mechanisms of Pathogenesis 424

III. Mutations in Cx26 Lead to Nonsyndromic

Deafness 42 8

IV. Implications of Cx26 Mutations fo r

Hearing 43 4

V. Mutations in Cx31 Lead to Autosoma lDominant Erythrokeratodermia Variabilis o r

Deafness 43 8VI. Mutations in Cx32 Lead to an Inherited Peripheral

Neuropathy 439VII. The Clinical Manifestations of CMTX 442

VIII. Cx32 Expression in Schwann Cells and Pathogenesis o fCMTX 443

IX. Mutations in Cx43 Were Found in a Few Patients withVisceroatrial Heterotaxia 44 8

X. Mutations in Cx46 and Cx50 Lead toCataracts 449

XI. Candidate Diseases for Other Connexins 450References 453

CHAPTER 21 Trafficking and Targeting of Connexin32 Mutation sto Gap Junctions in Charcot-Marie-ToothX-Linked DiseasePatricia E. M. Martin and W. Howard Evan s

I. Introduction 46 1II. Classification of Mutations in CMT-X 46 5

III. Mechanisms Leading to the Intracellular Trapping o fMutant Protein 47 3

IV. Gap Junction Targeting Determinants 47 4V. Mutations in Other Connexins and Disease 476

VI. Concluding Remarks 47 7References 477

CHAPTER 22 Molecular Basis of Deafness Due to Mutations in th eConnexin26 Gene (GJB2 )Xavier Estivill and Raquel Rabione t

I. Anatomic, Mechanical, and Neural Basis of Hearing 484II. Epidemiological Basis of Deafness 486

III. Defining the Genes of Deafness 48 8IV. Identification of GJB2 (Connexin26) (DFNBI) as a Gen e

Responsible for Deafness 490V. Connexin26 Gene Structure, Expression, and Function 49 2

VI. Deafness Mutations in GJB2 495VII. 35delG, a Frequent Mutation in the Genera l

Population 498VIII. GJB2 Mutation Analysis 499

IX. What Does Connexin26 Have to Do with Hearing? 50 0X. Other Connexins Involved in Deafness 50 1

XI. Clinical Consequences of GJB2 Mutations an dTherapeutic Implications 502References 504

CHAPTER 23 "Negative" Physiology: What Connexin-Deficien tMice Reveal about the Functional Roles of Individua lGap Junction Protein sD. C. Spray, T. Kojima, E. Scemes, S. O. Suadicani, Y. Gao ,S . Zhao, and A. Fort

I. Introduction 509II. Communication Compartments and Genetic Alteration s

Associated with Connexin Dysfunction 510

III. Connexin26 : Human Deafness, the Mouse Placenta, an dMechanisms of Autosomal Dominance 51 3

IV. Connexin32 : A Critical Component of IntracellularSignaling in Myelinating Schwann Cells and ofIntercellular Signaling in the Liver 51 4

V. Talk and Crosstalk in Brain Communicatio nCompartments 51 9

VI. Cx43, Cx46, and Cx50 in the Lens 520VII. Targeted Disruption of Connexin43 an d

Connexin40 Gene Expression: What Happens to th eHeart? 523

VIII. Conclusions 526References 52 7

CHAPTER 24 Role of Gap Junctions in Cellular Growth Control andNeoplasia: Evidence and MechanismsRandall J. Ruch

I. Introduction 53 5II. Evidence That GJIC Regulates Cellular Growth and Is

Involved in Neoplasia 53 7III. Mechanism of GJIC-Mediated Growth Control 545IV. Is GJIC Necessary, or Do Connexins Regulate Growth

Independently of GJIC? 546References 548

CHAPTER 25 Gap Junctions in Inflammatory Responses :Connexins, Regulation, and Possible Functional Role sJuan C. Säez, Roberto Araya, Maria C. Braves, Miguel Concha,Jorge E. Contreras, Eliseo A. Eugenin, Agustin D. Martinez,Francis Palisson, and Manuel A. Sepulveda

I. The Inflammatory Response : A Brief Introduction 555II. Steps of the Inflammatory Response That Induce Gap

Junction Changes 55 6III. Putative Mechanisms That Regulate Gap Junctions i n

Local Cells during an Inflammatory Process 55 7IV. Gap Junctional Communication between Cells of th e

Immune System 56 4V. Functional Consequences of Changes in Ga p

Junctional Communication during InflammatoryResponses 569References 572

CHAPTER 26 Cx43 (a,) Gap Junctions in Cardiac Development an dDiseas eRobert G. Gourdie and Cecilia W. Lo

I. Introduction 58 1II. a l and the Working Myocardium : Expression i n

Mammals and Nonmammals 582

III. a l and Other "Cardiac" Connexins 58 4

IV. a l Connexin Expression in Developing CardiacMuscle 585

V. Regulation of a l Distribution in the MyocyteSarcolemma in the Developing and Disease dHeart 586

VI. A Role for a l Connexin in Heart Development 58 8VII. Gap Junctions and the Modulation of Cardiac Neura l

Crest Migration 59 1VIII. Crest Abundance and Development of th e

Myocardium 59 3IX. The Role of a l Connexins in Working Myocytes 594X. a l Connexin Perturbation and Congenital Cardiac

Defects 595XI. Speculations 596

References 59 7

CHAPTER 27 Gap Junctional Communication in the Failing HeartWalmor C. De Mello

I. Calcium Overload and Healing Over in HeartFailure 604

II. Junctional Conductance and (3-Adrenergic Recepto rActivation in the Failing Heart 609

III. Renin-Angiotensin System and Heart Cel lCommunication 61 1

IV. Conclusion 62 0References 62 1

CHAPTER 28 Gap Junctions Are Specifically Disrupted byTrypanosoma cruzi InfectionRegina C. S. Goldenberg, Andrea Gonsalves,and Antonio C. Campos de Carvalh o

I. Introduction 625II. The Time Course of Uncoupling 626

III. Uncoupling Is Not Cell-, Connexin-, or Parasite -Specific 62 9

IV. T. cruzi Infection Specifically Disrupts Gap Junctio nCommunication in MDCK Cultures 630References 634

Index 635