Connexin Methods and Protocols

ISBN-13: 9780896036581 / Angielski / Twarda / 2000 / 491 str.

Roberto Bruzzone; Christian Giaume

Opis

Direct cell cell communication is a common property of multicellular organisms that is achieved through membrane channels which are organized in gap junctions. The protein subunits of these intercellular channels, the connexins, form a multigene family that has been investigated in great detail in recent years. It has now become clear that, in different tissues, connexins speak several languages that control specific cellular functions. This progress has been made possible by the availability of new molecular tools and the improvement of basic techniques for the study of membrane channels, as well as by the use of genetic approaches to study protein function in vivo. More important, connexins have gained visibility because mutations in some connexin genes have been found to be linked to human genetic disorders. Connexin Methods and Protocols presents in detail a collection of te- niques currently used to study the cellular and molecular biology of connexins and their physiological properties. The field of gap junctions and connexin research has always been characterized by a multidisciplinary approach c- bining morphology, biochemistry, biophysics, and cellular and molecular biology. This book provides a series of cutting-edge protocols and includes a large spectrum of practical methods that are available to investigate the fu- tion of connexin channels. Connexin Methods and Protocols is divided into three main parts."

Recenzje

"This book will be invaluable to those in the connexin field and a fascinating read and useful text to those new to this area of research." -Molecular Biotechnology

Spis treści

Part I. Tools to Study Connexins. Investigation of Connexin Gene Expression Patterns by In Situ Hybridization Techniques, Magali Théveniau-Ruissy, Sébastien Alcoléa, Irene Marics, Daniel Gros, Antoon F. M. Moorman, and Wouter H. Lamers. Sodium Dodecyl Sulfate-Freeze-Fracture Immunolabeling of Gap Junctions, Irene Dunia, Michel Recouvreur, Pierre Nicolas, Nalin M. Kumar, Hans Bloemendal, and E. Lucio Benedetti. Purification of Gap Junctions, Gina E. Sosinsky and Guy A. Perkins. Culturing of Mammalian Cells Expressing Recombinant Connexins and Two-Dimensional Crystallization of the Isolated Gap Junctions, Mark Yeager and Vinzenz M. Unger. Connexins/Connexons: Cell-Free Expression, Matthias M. Falk. Biochemical Analysis of Connexon Assembly, Judy K. VanSlyke and Linda S. Musil. Expression and Imaging of Connexin-GFP Chimeras in Live Mammalian Cells, Dale W. Laird, Karen Jordan, and Qing Shao. Analysis of Connexin Expression in Brain Slices by Single-Cell Reverse Transcriptase Polymerase Chain Reaction, Laurent Venance. Use of Retroviruses to Express Connexins, Jean X. Jiang. Spatiotemporal Depletion of Connexins Using Antisense Oligonucleotides, Colin R. Green, Lee yong Law, Jun Sheng Lin, and David L. Becker. Transfection and Expression of Exogenous Connexins in Mammalian Cells, Dieter Manthey and Klaus Willecke. Part II. Assays for Function. Assaying the Molecular Permeability of Connexin Channels, Paolo Meda. Applying the Xenopus Oocyte Expression System to the Analysis of Gap Junction Proteins, I. Martha Skerrett, Mary Merritt, Lan Zhou, Hui Zhu, FengLi Cao, Joseph F. Smith, and Bruce J. Nicholson. Mutagenesis to Study Channel Structure, Gerhard Dahl and Arnold Pfahnl. Dual Patch Clamp, Harold V.M. Van Rijen, Ronald Wilders, Martin B. Rook, and Habo J. Jongsma. Determining Ionic Permeabilities of Gap Junction Channels, Richard D. Veenstra. Fluorescence Recovery After Photobleaching, Jean Délèze, Bruno Delage, Olfa Hentati-Ksibi, Franck Verrecchia, and JeanClaude Hervé. Capture of Transjunctional Metabolites, Gary S. Goldberg and Paul D. Lampe. Part III. Physiology and Biology of Connexins. The Study of Connexin Hemichannels (Connexons) in Xenopus Oocytes, E. Brady Trexler and Vytas K. Verselis. Exploring Hemichannel Permeability In Vitro, Andrew L. Harris and Carville G. Bevans. Inducing De Novo Formation of Gap Juntion Channels, Feliksas F. Bukauskas. Recording and Analysis of Putative Direct Electrical Interactions in the Mammalian Brain, Taufik A. Valiante, Jose L. Perez Velazquez, and Peter L. Carlen. Intercellular Calcium Signaling and Flash Photolysis of Caged Compounds: A Sensitive Method to Evaluate Gap Junctional Coupling, Luc Leybaert and Michael J. Sanderson. Biochemical Analysis of Connexin Phosphorylation, Bonnie J. Warn-Cramer, Wendy E. Kurata, and Alan F. Lau. How to Close a Gap Junction Channel: Efficacies and Potencies of Uncoupling Agents, Renato Rozental, Miduturu Srinivas, and David C. Spray. Index.

Blurb

An essential element in direct cell-cell communication between virtually all cell types in vertebrates, connexins have been recently linked to an increasing number of human genetic diseases. In Connexin Methods and Protocols, well established investigators describe in detail their best techniques for studying the cellular and molecular biology of connexins, as well as their physiological properties. These forefront methods are specifically designed to investigate the life of connexins from biosynthesis to assembly into gap junction channels, and also include powerful assays to perform functional studies on these routes of cell-cell communication. To ensure experimental success, each time-tested technique contains detailed step-by-step instructions, as well as tips on how to avoid pitfalls. Also discussed are the many specific
theoretical aspects of connexin biology and physiology.
Cutting-edge and highly practical, Connexin Methods and Protocols offers laboratory researchers a comprehensive collection of proven methods for studying the role of connexins in intercellular communication and human genetic disorders, as well as finding promising new therapeutics for those disorders.