Semaphorin Signaling

This volume arranged into three sections describes biochemical, in vitro, and in vivo protocols on Semaphorins. Chapters focus on approaches that would allow the novice to study Semaphorins and employ robust assays to characterize mechanisms of action. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Semaphorin Signaling: Methods and Protocols aims to ensure successful results in the further study of this vital field.

Includes cutting-edge methods and protocols Provides step-by-step detail essential for reproducible results Contains key notes and implementation advice from the experts Includes supplementary material: sn.pub/extras

Klappentext

This volume arranged into three sections describes biochemical, in vitro, and in vivo protocols on Semaphorins. Chapters focus on approaches that would allow the novice to study Semaphorins and employ robust assays to characterize mechanisms of action. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Authoritative and cutting-edge, Semaphorin Signaling: Methods and Protocols aims to ensure successful results in the further study of this vital field.

Inhalt

Semaphorins and their Signaling Mechanisms.- Isolating Fc-Tagged SEMA4D Recombinant Protein from 293FT Cells.- Expression and Purification of Class 7 Semaphorin and its Plexinc1 Receptor Using Baculovirus-Mediated Mammalian Cell Gene Transduction.- Immunoaffinity Purification of the Glycosylated Extracellular Fragment of Mouse Plexin A2 Produced in a Mammalian Expression Systems.- Plate-Based Assay for Measuring Direct Semaphorin-Neuropilin Interactions.- Characterizing Plexin Gtpase Interactions Using Gel Filtration, Surface Plasmon Resonance Spectrometry and Isothermal Titration Calorimetry.- In Vitro Assay for the Rap Gtpase Activating Protein Activity of the Purified Cytoplasmic Domain of Plexin.- Characterizing F-Actin Disassembly Induced by the Semaphorin Signaling Component.- Characterizing Erbb-2-Mediated Tyrosine Phosphorylation and Activation of Plexins.- Characterizing PKA-Mediated Phosphorylation of Plexin Using Purified Proteins.- Using Heterologous Cos-7 Cells to Identify Semaphorin Signaling Components.- Analysis of Semaphorin-Induced Growth Cone Collapse and Axon Growth Inhibition.- Using Rotary Shadow Electron Microscopy to Characterize Semaphorin-Mediated Growth Cone Collapse.- An Electrical Impedance-Based Method for Quantitative Real-Time Analysis of Semaphorin-Elicited Endothelial Cell Collapse.- Regulation of Cortical Dendrite Morphology and Spine Organization by Secreted Semaphorins A Primary Culture Approach.- Characterizing Semaphorin Signaling Using Isolated Neurons in Culture.- Performing Axon Orientation Assays With Secreted Semaphorins And Other Guidance Cues.- Assays to Examine Transmembrane Semaphorin Function In Vitro.- Micro-CALI to Study Localized Roles of the Semaphorin Signaling Component CRMP in Axon Growth.- Visualizing and Characterizing Semaphorin Endocytic Events Using Quantum Dot Conjugated Proteins.- Visualization of Clathrin-Mediated Endocytosis during Semaphorin-Guided Axonal Growth.- Tracking Differential Endocytosis and Trafficking of Semaphorin Receptor Complexes in Responding Nerve Growth Cones.- Antibody-Feeding Assay: A Method to Track the Internalization of Neuropilin-1 and Other Cell Surface Receptors.- Photolithography-Based Substrate Microfabrication for Patterning Semaphorin 3A to Study Neuronal Development.- Characterization of Semaphorin 6A-Mediated Effects on Angiogenesis through Regulation of VEGF Signaling.- Studying the Effects of Semaphorins on Oligodendrocyte Lineage Cells.- Characterizing Semaphorin-Mediated Immune Responses Using an Antigen-Presentation Assay.- Podocyte Shape Regulation by Semaphorin 3A and MICAL-1.- In Vivo and In Vitro Knockdown Approachesin the Avian Embryo as a Means to Study Semaphorin Signaling.- Assaying the Action of Secreted Semaphorins on Vascular Permeability.- Characterization of the Effects of Semaphorin 4D Signaling on Angiogenesis.- Characterizing Semaphorin-Mediated Effects on Sensory and Motor Axon Pathfinding and Connectivity during Embryonic Development.- Experimental Approaches for Studying Semaphorin Signals in Tumor Growth and Metastasis in Mouse Models.- Characterizing Semaphorin Signaling In Vivo Using C. Elegans.