Ras Signaling: Methods and Protocols

ISBN-13: 9781493960545 / Angielski / Miękka / 2016 / 425 str.

Lorenza Trabalzini; Saverio Francesco Retta

Opis

Featuring experimental approaches that shed light on the complexity of Ras GTPase biological functions, Ras Signaling: Methods and Protocols contains general overviews and detailed applications of both well-established and recently developed research techniques, including biochemical, biophysical, molecular biology, genetic and behavioral approaches, advanced high resolution fluorescence and electron microscopy imaging and "omics" technologies. Through this, the detailed volume provides information on expression, post-translational modifications, subcellular localization and dynamics, regulatory mechanisms of upstream and downstream signaling pathways and ultimately, biological activities and functions of Ras GTPases in different model systems, including high and low eukaryotic organisms. Written in the highly successful Methods in Molecular Biology series format, chapters include brief introductions, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols and tips on troubleshooting and avoiding known pitfalls.Wide-ranging and authoritative, Ras Signaling: Methods and Protocols serves as an aid for investigators of different backgrounds and interests related to the multiple physiological and pathological functions of the large superfamily of Ras GTPases.

Spis treści

1. The Ras Superfamily of Small GTPases: The Unlocked Secrets

            Luca Goitre, Eliana Trapani, Lorenza Trabalzini, and Saverio Francesco Retta

2. A Novel Method for the Production of Fully-Modified K-Ras 4B

            Tanmay S. Chavan, James O. Meyer, Luke Chisholm, Malgorzata Dobosz, and Vadim Gaponenko

3. Metabolic Labeling of Ras with Tritiated Palmitate to Monitor Palmitoylation and Depalmitoylation

            Frederick D. Tsai, Joseph P. Wynne, Ian M. Ahearn, and Mark R. Philips

4. Ubiquitination of Rac1 by Inhibitors of Apoptosis (IAPs)

            Tripat Kaur Oberoi-Khanuja and Krishnaraj Rajalingam

5. Ras GTPases are Both Regulators and Effectors of Redox Agents

            Elisa Ferro, Luca Goitre, Eva Baldini, Saverio Francesco Retta, and Lorenza Trabalzini

6. Biophysical and Proteomic Characterization Strategies for Cysteine Modifications in Ras GTPases

            G. Aaron Hobbs, Harsha P. Gunawardena, and Sharon L. Campbell

7. Use of the Yeast Two-Hybrid Technology to Isolate Molecular Interactions of Ras GTPases

            Elisa Ferro, Eva Baldini, and Lorenza Trabalzini

8. Screening for MAPK Modulators Using an In-Cell Western Assay

            Simon Schnaiter, Beatrix Fürst, Johannes Neu, Frigyes Wáczek, László Őrfi, György Kéri, Lukas A. Huber, and Winfried Wunderlich

9. Behavioral Methods for the Study of the Ras-ERK Pathway in Memory Formation and Consolidation: Passive Avoidance and Novel Object Recognition Tests

            Raffaele d’Isa, Riccardo Brambilla, and Stefania Fasano

10. Functional Phosphoproteomics of Oncogenic KRAS Signaling

            Putty-Reddy Sudhir and Jeou-Yuan Chen

11. Pull-Down Assay for Analysis of Integrin-Mediated Activation of Rap Proteins in Adherent Platelets

            Gianni Francesco Guidetti and Mauro Torti

12. Combined Pulldown and Time-Lapse Microscopy Studies for Determining the Role of Rap1 in the Crosstalk between Integrins and Cadherins

            Luca Goitre, Alessandro Morina, and Saverio Francesco Retta

13. Fluorescence Microscopy Study of Rap1 Subcellular Localization

            Luca Goitre, Valentina Cutano, and Saverio Francesco Retta

14. An In Vitro System to Evaluate the Scaffold Function of the RalA Effector Protein RalBP1

            David F. Kashatus

15. Analysis of the Rit Subfamily GTPase-Mediated Signaling and Neuronal Differentiation and Survival

            Geng-Xian Shi and Catherine N. Kaminski

16. Immunofluorescence Methods in Studies of the GTPase RAN and Its Effectors in Interphase and in Mitotic Cells

            Giulia Guarguaglini, Valeria de Turris, and Patrizia Lavia

17. High Resolution Scanning Electron Microscopy for the Imaging of Nuclear Pore Complexes and Ran-Mediated Transport

            Lihi Shaulov, Boris Fichtman, and Amnon Harel

18. Effector Recruitment Method to Study Spatially Regulated Activation of Ras and Rho GTPases

            Lauren P. Huff, Molly J. DeCristo, and Adrienne D. Cox

19. Real-Time Visualization and Quantification of Native Ras Activation in Single Living Cells

            Christoph Biskup and Ignacio Rubio

20. Nanoclustering and Heterogeneous Membrane Diffusion of Ras Studied by FRAP and RICS Analysis

            Camilo Guzmán, Maja Šolman, and Daniel Abankwa

21. Analyzing the Roles of Rho GTPases in Cancer Cell Migration with a Live Cell Imaging 3D-Morphology-Based Assay

            Audrey Colomba and Anne J. Ridley

22. Analysis of Rho-GTPase-Induced Localization of Nanoscale Adhesions Using Fluorescence Nanoscopy

            Annica K.B. Gad, Daniel Rönnlund, Jerker Widengren, and Pontus Aspenström

 

23. Yeast as a Model for Ras Signaling

            Renata Tisi, Fiorella Belotti, and Enzo Martegani

24. Methods to Study the Ras2 Protein Activation State and the Subcellular Localization of Ras-GTP in Saccharomyces cerevisiae

            Sonia Colombo and Enzo Martegani

25. RAS  Proteins Signaling in the Early Metazoan Dictyostelium discoideum

            Enrico Bracco and Barbara Pergolizzi