Rho GTPases: Methods and Protocols

Part I. History and Phylogenesis

1. Historical Overview of Rho GTPases

Anne J. Ridley

2. Rho GTPases: Deciphering the Evolutionary History of a Complex Protein Family

Marek Eliás and Vladimír Klimes

Part II. Biochemistry of Rho GTPases

3. Biochemical Assays to Characterize Rho GTPases

Mamta Jaiswal, Badri N. Dubey, Katja T. Koessmeier, Lothar Gremer, and Mohammad R. Ahmadian

4. How to Analyze Bacterial Toxins Targeting Rho GTPases

Heike Bielek and Gudula Schmidt

5. Assessing Ubiquitylation of Rho GTPases in Mammalian Cells

Anne Doye, Amel Mettouchi, and Emmanuel Lemichez

6. Posttranslational Lipid Modification of Rho Family Small GTPases

Natalia Mitin, Patrick J. Roberts Emily J. Chenette, and Channing J. Der

7. Analysis of the Role of RhoGDI1 and Isoprenylation in the Degradation of Rho GTPases

Etienne Boulter and Rafael Garcia-Mata

8. A Quantitative Fluorometric Approach for Measuring the Interaction of RhoGDI with Membranes and Rho GTPases

Jared Johnson, Richard A. Cerione, and Jon W. Erickson

Part III. Functional assays

9. Rho GTPases and Cancer Cell Transendothelial Migration

Nicolas Reymond, Philippe Riou, and Anne J. Ridley

10. Quantitative and Robust Assay to Measure Cell-cell Contact Assembly and Maintenance

Sébastien Nola, Jennifer C. Erasmus, and Vania M. M. Braga

11. Rho GTPase Knockout Induction in Primary Keratinocytes from Adult Mice

Esben Pedersen, Astrid Basse, Tine Lefever, Karine Peyrollier, and Cord Brakebusch

12. Rho GTPase Techniques in Osteoclastogenesis

Roland Leung and Michael Glogauer

13. Assessment of Rho GTPase Signaling During Neurite Outgrowth

Daniel Feltrin and Olivier Pertz

14. Assesment of the Role for Rho Family GTPases in NADPH Oxidase Activation

Kei Miyano and Hideki Sumimoto

Part IV. Imaging and high throughput methods

15. Multiplex Imaging of Rho GTPase Activities in Living Cells

Désirée Spiering and Louis Hodgson

16. FRET-Based Imaging of Rac and Cdc42 Activation During Fc-Receptor Mediated Phagocytosis in Macrophages

Adam D. Hoppe

17. High Throughput Flow Cytometry Bead-Based Multiplex Assay for Identification of Rho GTPase Inhibitors

Zurab Surviladze Susan M. Young, and Larry A. Sklar

18. Functional Analysis of RhoGTPase Activation and Inhibition in a Bead-Based Miniaturized Format

Michael Schmohl, Stefanie Rimmele, Peter Gierschik, Thomas O. Joos, and Nicole Schneiderhan-Marra

19. Use of Phage Display For the Identification of Molecular Sensors Specific for Activated Rho

Patrick Chinestra, Isabelle Lajoie-Mazenc, Jean-Charles Faye, and Gilles Favre

20. Identification of New Interacting Partners for Atypical Rho-GTPases: a SILAC-Based Approach

Laura Montani, Ana Filipa Domingues, and João Bettencourt Relvas

Part V. Rho GTPases in Non-Mammalian Model Organisms

21. Using Zebrafish for Studying Rho GTPases Signaling in Vivo

Shizhen Zhu and Boon Chuan Low

22. Analysis of Rho GTPase Function in Axon Pathfinding Using Caenorhabditis elegans

Jamie K. Alan and Erik A. Lundquist

23. Protocol for ex vivo Incubation of Drosophila Primary Post-Embryonic Haemocytes for Real-Time Analyses

Christopher J. Sampson and Michael J. Williams

24. Analysis of Rho GTPase Activation in Saccharomyces cerevisiae

Gary Eitzen and Michael R. Logan

25. Assaying Rho GTPase-Dependent Processes in Dictyostelium discoideum

Huajiang Xiong and Francisco Rivero

26. FRAP-Based Analysis of Rho GTPase-dependent Polar Exocytosis in Pollen Tubes

An Yan and Zhenbiao Yang