Contents Preface Contributors Part I An Overview of Methods for Analyzing Vesicular Trafficking 1 Vesicular Trafficking: Molecular Tools and Targets Elena Vassilieva and Asma Nusrat 2 Pharmacological Inhibition of Endocytic Pathways. Is It Specific Enough to be Useful? Andrei Ivanov Part II Cell-free and Biochemical Assays of Exocytosis and Endocytosis 3 In Vitro Assays to Measure SNARE-Mediated Vesicle Fusion Susanne Kreye, Jörg Malsam, and Thomas H. Söllner 4 Imaging Ca2+-Triggered Exocytosis of Single Secretory Granules on Plasma Membrane Lawns from Neuroendocrine Cells. Thorsten Lang 5 FRAP Analysis of Secretory Granule Lipids and Proteins in the Sea Urchin Egg Julian L. Wong and Gary M. Wessel 6 A Cell-Free Assay for Endocytosis of E-cadherin Toshiaki Sakisaka and Yoshimi Takai 7 Cell-Surface Biotinylation to Study Endocytosis and Recycling of Occludin Noriyuki Nishimura and Takuya Sasaki 8 Fractionation of Subcellular Membrane Vesicles of Epithelial and Non-epithelial Cells by OptiPrepTM Density Gradient Ultracentrifugation Xuhang Li and Mark Donowitz 9 Identification and Characterization of Interacting Partners of Rab GTPases by Yeast Two-Hybrid Analyses Mark Kail and Angelika Barnekow Part III Analysis of Endocytosis and Exocytosis in Different Cell Systems 10 High-Throughput Analysis of the Dynamics of Recycling Cell Surface Proteins Roland Govers, David E. James, and Adelle C. F. Coster 11 Studying Phagocytosis by Live-Cell Scintillation Proximity Assay Walter Stockinger and Axel Nohturfft 12 Transcytosis of Polymeric Immunoglobulin A in Polarized Madin-Darby Canine Kidney Cells Asli Oztan, Christine Rondanino, and Gerard Apodaca 13 The Use of SyntaxinChimeras to Study Polarized Protein Trafficking in Epithelial Cells Martin B.A. ter Beest 14 Targeting the Epithelial SNARE Machinery by Bacterial Neurotoxins Veronique Proux-Gillardeaux and Thierry Galli 15 Exocytosis of Endothelial Cells is Regulated by N-ethylmaleimide Sensitive Factor Munekazu Yamakuchi, Marcella Ferlito, Craig N. Morrell, Kenji Matsushita, Craig A. Fletcher, Wangsen Cao, and Charles J. Lowenstein 16 Molecular Dissection of HCl Secretion in Gastric Parietal Cells Using Streptolysin O Permeabilization Xia Ding, Fang Wu, Zhen Guo, and Xuebiao Yao 17 Addressing Membrane Protein Topology Using the Fluorescence Protease Protection (FPP) Assay Holger Lorenz, Dale W. Hailey, and Jennifer Lippincott-Schwartz 18 Visualizing Clathrin-Mediated IgE Receptor Internalization by Electron and Atomic Force Microscopy Alan R. Burns, Janet M. Oliver, Janet R. Pfeiffer, and Bridget S. Wilson 19 Analyzing Endosomes in Non-Sectioned Cells by Transmission Electron Microscopy Willem Stoorvogel 20 Imaging Exocytosis of Single Insulin Secretory Granules with TIRF Microscopy Shinya Nagamatsu and Mica Ohara-Imaizumi 21 Reconstitution of Depolarization and Ca2+-Evoked Secretion in Xenopus Oocytes Monitored by Membrane Capacitance Roy Cohen, Bernhard M. Schmitt, and Daphne Atlas 22 Low-Noise Recording of Single Vesicle Capacitance Steps in Cell-Attached Patches Vitaly Klyachko, Zhenjie Zhang, and Meyer Jackson 23 Good Practices in Single Cell Amperometry J. David Machado, Mónica S. Montesinos, and Ricardo Borges 24 Analysis of Single-Vesicle Exocytotic Events Recorded by Amperometry Eugene V. Mosharov Part IV. Analysis of Exocytosis and Endocytosis in Model Organisms 25 Application of RNAi Technology and Fluorescent Protein Markers to Study Membrane Traffic in C. elegans
In this book, skilled experts provide the most up-to-date, step-by-step laboratory protocols for examining molecular machinery and biological functions of exocytosis and endocytosis in vitro and in vivo. The book is insightful to both newcomers and seasoned professionals. It offers a unique and highly practical guide to versatile laboratory tools developed to study various aspects of intracellular vesicle trafficking in simple model systems and living organisms.
Exocytosis and endocytosis remain among the most popular subjects in biology and biomedical sciences. Tremendous progress in understanding these complex intracellular processes has been achieved by employing a wide array of research tools ranging from classical biochemical methods to modern imaging techniques. In this book, skilled experts provide the most up-to-date, step-by-step laboratory protocols for examining molecular machinery and biological functions of exocytosis and endocytosis in vitro and in vivo. The volume presents detailed lists of tested reagents that really work, and notes on troubleshooting from leading laboratories in the field. Insightful to both newcomers and seasoned professionals, the book offers a unique and highly practical guide to versatile laboratory tools developed to study various aspects of intracellular vesicle trafficking in simple model systems and living organisms.