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 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
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.
1. Semaphorins and their Signaling Mechanisms
Laura T. Alto and Jonathan R. Terman
Part I: Biochemical Approaches
2. Isolating Fc-Tagged SEMA4D Recombinant Protein from 293FT Cells
Joanne Soong and Glynis A. Scott
3. Expression and Purification of Class 7 Semaphorin and its Plexinc1 Receptor Using Baculovirus-Mediated Mammalian Cell Gene Transduction
Xiaoyan Chen, Po-Han Chen, and Xiaolin He
4. Immunoaffinity Purification of the Glycosylated Extracellular Fragment of Mouse Plexin A2 Produced in a Mammalian Expression System
Terukazu Nogi, Emiko Mihara, Norihisa Yasui, and Junichi Takagi
5. Plate-Based Assay for Measuring Direct Semaphorin-Neuropilin Interactions
Matthew W. Parker and Craig W. Vander Kooi
6. Characterizing Plexin Gtpase Interactions Using Gel Filtration, Surface Plasmon Resonance Spectrometry and Isothermal Titration Calorimetry
Jeannine Muller-Greven, Soonjeung Kim, Prasanta K. Hota, Yufeng Tong, Susmita Borthakur, and Matthias Buck
7. In Vitro Assay for the Rap Gtpase Activating Protein Activity of the Purified Cytoplasmic Domain of Plexin
Heath G. Pascoe, Yuxiao Wang, and Xuewu Zhang
8. Characterizing F-Actin Disassembly Induced by the Semaphorin Signaling Component Mical
Jimok Yoon, Ruei-Jiun Hung, and Jonathan R. Terman
9. Characterizing Erbb-2-Mediated Tyrosine Phosphorylation and Activation of Plexins
Thomas Worzfeld and Jakub M. Swiercz
10. Characterizing PKA-Mediated Phosphorylation of Plexin Using Purified Proteins
Taehong Yang and Jonathan R. Terman
Part II: In Vitro Approaches
11. Using Heterologous Cos-7 Cells to Identify Semaphorin Signaling Components Atsuko Sakurai, Colleen L. Doçi, and J. Silvio Gutkind
12. Analysis of Semaphorin-Induced Growth Cone Collapse and Axon Growth Inhibition
Lionel Meyer, Andrius Kaselis, Saulius Satkauskas,
and Dominique Bagnard
13. Using Rotary Shadow Electron Microscopy to Characterize Semaphorin-Mediated Growth Cone Collapse
Paul C. Bridgman
14. An Electrical Impedance-Based Method for Quantitative Real-Time Analysis of Semaphorin-Elicited Endothelial Cell Collapse
Chiara Camillo, Noemi Gioelli, Federico Bussolin, and Guido Serini
15. Regulation of Cortical Dendrite Morphology and Spine Organization by Secreted Semaphorins – A Primary Culture Approach
Sheng-Shiang Peng and Tracy S. Tran
16. Characterizing Semaphorin Signaling Using Isolated Neurons in Culture
Sophie Chauvet, Erik Mire, and Fanny Mann
17. Performing Axon Orientation Assays With Secreted Semaphorins And Other Guidance Cues
Céline Delloye-Bourgeois, Frédéric Moret And Valérie Castellani
18. Assays to Examine Transmembrane Semaphorin Function In Vitro
Rachel Q. Liu, Kristen Browne, and Timothy P. O´Connor
19. Micro-CALI to Study Localized Roles of the Semaphorin Signaling Component CRMP in Axon Growth
Yoshio Goshima and Kohtaro Takei
20. Visualizing and Characterizing Semaphorin Endocytic Events Using Quantum Dot Conjugated Proteins
Ioana Carcea and Deanna L. Benson
21. Visualization of Clathrin-Mediated Endocytosis during Semaphorin-Guided Axonal Growth
Rurika Itofusa, Takuro Tojima, and Hiroyuki Kamiguchi
22. Tracking Differential Endocytosis and Trafficking of Semaphorin Receptor Complexes in Responding Nerve Growth Cones
Puneet Dang and Andrew J. W. Furley
23. Antibody-Feeding Assay: A Method to Track the Internalization of Neuropilin-1 and Other Cell Surface Receptors
Sabrina Rizzolio and Luca Tamagnone
24. Photolithography-Based Substrate Microfabrication for Patterning Semaphorin 3A to Study Neuronal Development
Maya Shelly, Seong-Il Lee, Giulia Suarato, Yizhi Meng, and Sophie Pautot
25. Characterization of Semaphorin 6A-Mediated Effects on Angiogenesis through Regulation of VEGF Signaling
Hidetaka Ohnuki and Giovanna Tosato
26. Studying the Effects of Semaphorins on Oligodendrocyte Lineage Cells
Yasir Ahmed Syed Sarah Ali Abdulla, and Mark. R. N. Kotter
27. Characterizing Semaphorin-Mediated Immune Responses Using an Antigen-Presentation Assay
Satoshi Nojima and Atsushi Kumanogoh
28. Podocyte Shape Regulation by Semaphorin 3A and MICAL-1
Alda Tufro
Part III: In Vivo Approaches
29. In Vivo and In Vitro Knockdown Approaches in the Avian Embryo as a Means to Study Semaphorin Signaling
Karli Montague, Sarah Guthrie, and Ivana Poparic
30. Assaying the Action of Secreted Semaphorins on Vascular Permeability
Lucas Treps and Julie Gavard
31. Characterization of the Effects of Semaphorin 4D Signaling on Angiogenesis
Hua Zhou, Ying-Hua Yang, and John R. Basile
32. Characterizing Semaphorin-Mediated Effects on Sensory and Motor Axon Pathfinding and Connectivity during Embryonic Development
Rosa Eva Huettl
and Andrea B. Huber
33. Experimental Approaches for Studying Semaphorin Signals in Tumor Growth and Metastasis in Mouse Models
Enrico Moiso, Massimo Accardo, and Luca Tamagnone
34. Characterizing Semaphorin Signaling In Vivo Using C. Elegans
Akira Nukazuka and Shin Takagi
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.
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.
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.
Inhaltsverzeichnis
1. Semaphorins and their Signaling Mechanisms
Laura T. Alto and Jonathan R. Terman
Part I: Biochemical Approaches
2. Isolating Fc-Tagged SEMA4D Recombinant Protein from 293FT Cells
Joanne Soong and Glynis A. Scott
3. Expression and Purification of Class 7 Semaphorin and its Plexinc1 Receptor Using Baculovirus-Mediated Mammalian Cell Gene Transduction
Xiaoyan Chen, Po-Han Chen, and Xiaolin He
4. Immunoaffinity Purification of the Glycosylated Extracellular Fragment of Mouse Plexin A2 Produced in a Mammalian Expression System
Terukazu Nogi, Emiko Mihara, Norihisa Yasui, and Junichi Takagi
5. Plate-Based Assay for Measuring Direct Semaphorin-Neuropilin Interactions
Matthew W. Parker and Craig W. Vander Kooi
6. Characterizing Plexin Gtpase Interactions Using Gel Filtration, Surface Plasmon Resonance Spectrometry and Isothermal Titration Calorimetry
Jeannine Muller-Greven, Soonjeung Kim, Prasanta K. Hota, Yufeng Tong, Susmita Borthakur, and Matthias Buck
7. In Vitro Assay for the Rap Gtpase Activating Protein Activity of the Purified Cytoplasmic Domain of Plexin
Heath G. Pascoe, Yuxiao Wang, and Xuewu Zhang
8. Characterizing F-Actin Disassembly Induced by the Semaphorin Signaling Component Mical
Jimok Yoon, Ruei-Jiun Hung, and Jonathan R. Terman
9. Characterizing Erbb-2-Mediated Tyrosine Phosphorylation and Activation of Plexins
Thomas Worzfeld and Jakub M. Swiercz
10. Characterizing PKA-Mediated Phosphorylation of Plexin Using Purified Proteins
Taehong Yang and Jonathan R. Terman
Part II: In Vitro Approaches
11. Using Heterologous Cos-7 Cells to Identify Semaphorin Signaling Components Atsuko Sakurai, Colleen L. Doçi, and J. Silvio Gutkind
12. Analysis of Semaphorin-Induced Growth Cone Collapse and Axon Growth Inhibition
Lionel Meyer, Andrius Kaselis, Saulius Satkauskas,
and Dominique Bagnard
13. Using Rotary Shadow Electron Microscopy to Characterize Semaphorin-Mediated Growth Cone Collapse
Paul C. Bridgman
14. An Electrical Impedance-Based Method for Quantitative Real-Time Analysis of Semaphorin-Elicited Endothelial Cell Collapse
Chiara Camillo, Noemi Gioelli, Federico Bussolin, and Guido Serini
15. Regulation of Cortical Dendrite Morphology and Spine Organization by Secreted Semaphorins - A Primary Culture Approach
Sheng-Shiang Peng and Tracy S. Tran
16. Characterizing Semaphorin Signaling Using Isolated Neurons in Culture
Sophie Chauvet, Erik Mire, and Fanny Mann
17. Performing Axon Orientation Assays With Secreted Semaphorins And Other Guidance Cues
Céline Delloye-Bourgeois, Frédéric Moret And Valérie Castellani
18. Assays to Examine Transmembrane Semaphorin Function In Vitro
Rachel Q. Liu, Kristen Browne, and Timothy P. O'Connor
19. Micro-CALI to Study Localized Roles of the Semaphorin Signaling Component CRMP in Axon Growth
Yoshio Goshima and Kohtaro Takei
20. Visualizing and Characterizing Semaphorin Endocytic Events Using Quantum Dot Conjugated Proteins
Ioana Carcea and Deanna L. Benson
21. Visualization of Clathrin-Mediated Endocytosis during Semaphorin-Guided Axonal Growth
Rurika Itofusa, Takuro Tojima, and Hiroyuki Kamiguchi
22. Tracking Differential Endocytosis and Trafficking of Semaphorin Receptor Complexes in Responding Nerve Growth Cones
Puneet Dang and Andrew J. W. Furley
23. Antibody-Feeding Assay: A Method to Track the Internalization of Neuropilin-1 and Other Cell Surface Receptors
Sabrina Rizzolio and Luca Tamagnone
24. Photolithography-Based Substrate Microfabrication for Patterning Semaphorin 3A to Study Neuronal Development
Maya Shelly, Seong-Il Lee, Giulia Suarato, Yizhi Meng, and Sophie Pautot
25. Characterization of Semaphorin 6A-Mediated Effects on Angiogenesis through Regulation of VEGF Signaling
Hidetaka Ohnuki and Giovanna Tosato
26. Studying the Effects of Semaphorins on Oligodendrocyte Lineage Cells
Yasir Ahmed Syed Sarah Ali Abdulla, and Mark. R. N. Kotter
27. Characterizing Semaphorin-Mediated Immune Responses Using an Antigen-Presentation Assay
Satoshi Nojima and Atsushi Kumanogoh
28. Podocyte Shape Regulation by Semaphorin 3A and MICAL-1
Alda Tufro
Part III: In Vivo Approaches
29. In Vivo and In Vitro Knockdown Approaches in the Avian Embryo as a Means to Study Semaphorin Signaling
Karli Montague, Sarah Guthrie, and Ivana Poparic
30. Assaying the Action of Secreted Semaphorins on Vascular Permeability
Lucas Treps and Julie Gavard
31. Characterization of the Effects of Semaphorin 4D Signaling on Angiogenesis
Hua Zhou, Ying-Hua Yang, and John R. Basile
32. Characterizing Semaphorin-Mediated Effects on Sensory and Motor Axon Pathfinding and Connectivity during Embryonic Development
Rosa Eva Huettl
and Andrea B. Huber
33. Experimental Approaches for Studying Semaphorin Signals in Tumor Growth and Metastasis in Mouse Models
Enrico Moiso, Massimo Accardo, and Luca Tamagnone
34. Characterizing Semaphorin Signaling In Vivo Using C. Elegans
Akira Nukazuka and Shin Takagi
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.n
nnnAuthoritative and cutting-edge, Semaphorin Signaling: Methods and Protocols aims to ensure successful results in the further study of this vital field.nn
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
