Abstract
The model organism D. discoideum is well suited to investigate basic questions of molecular and cell biology, particularly those related to the structure, regulation, and dynamics of the cytoskeleton, signal transduction, cell–cell adhesion, and development. D. discoideum makes use of Rho-regulated signaling pathways to reorganize its cytoskeleton during chemotaxis, endocytosis, and cytokinesis. In this organism the Rho family encompasses 20 members, several belonging to the Rac subfamily, but there are no representatives of the Cdc42 and Rho subfamilies. Here we present protocols suitable for monitoring the actin polymerization response and the activation of Rac upon stimulation of aggregation competent cells with the chemoattractant cAMP.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Kessin, R.H. (2001) Dictyostelium – Evolution, cell biology, and the development of multicellularity. Cambridge Univ. Press, Cambridge.
Steinert, M., and Heuner, K. (2005) Dictyostelium as host model for pathogenesis. Cell. Microbiol 7, 307–314.
Eichinger, L., and Rivero, F. (2006) Dictyostelium discoideum protocols. Methods in molecular biology 346. Humana Press Inc, Totowa.
Vlahou, G., and Rivero, F. (2006) Rho GTPase signaling in Dictyostelium discoideum: insights from the genome. Eur J Cell Biol 85, 947–959.
Park, K.C., Rivero, F., Meili, R., Lee, S., Apone, F., and Firtel, R.A. (2004) Rac regulation of chemotaxis and morphogenesis in Dictyostelium. EMBO J 23, 4177–4189.
Han, J. W., Leeper, L., Rivero, F., and Chung, C.Y. (2006) Role of RacC for the regulation of WASP and phosphatidylinositol 3-kinase during chemotaxis of Dictyostelium. J Biol Chem 281, 35224–35234.
Rivero, F., Illenberger, D., Somesh, B.P., Dislich, H., Adam, N., and Meyer, A.K. (2002) Defects in cytokinesis, actin reorganization and the contractile vacuole in cells deficient in RhoGDI. EMBO J. 21, 4539–4549.
Somesh, B.P., Vlahou, G., Iijima, M., Insall, R.H., Devreotes, P., and Rivero, F. (2006) RacG regulates morphology, phagocytosis, and chemotaxis. Eukaryot Cell 5, 1648–1663.
Chung, C.Y., Lee, S., Briscoe, C., Ellsworth, C., and Firtel, R.A. (2000) Role of Rac in controlling the actin cytoskeleton and chemotaxis in motile cells. Proc. Natl. Acad. Sci. USA 97, 5225–5230.
Larochelle, D.A., Vithalani, K.K., and De Lozanne, A. (1996) A novel member of the Rho family of small GTP-binding proteins is specifically required for cytokinesis. J. Cell Biol. 133, 1321–1329.
Dumontier, M., Hocht, P., Mintert, U., and Faix, J. (2000) Rac1 GTPases control filopodia formation, cell motility, endocytosis, cytokinesis and development in Dictyostelium. J. Cell Sci. 113, 2253–2265.
Somesh, B.P., Neffgen, C., Iijima, M., Devreotes, P., and Rivero, F. (2006) Dictyostelium RacH regulates endocytic vesicular trafficking and is required for localization of vacuolin. Traffic 7, 1194–1212.
Lee, E., Seastone, D.J., Harris, E., Cardelli, J.A., and Knecht, D.A. (2003) RacB regulates cytoskeletal function in Dictyostelium spp. Euk. Cell 2, 474–485.
Rivero, F., Albrecht, R., Dislich, H., Bracco, E., Graciotti, L., Bozzaro, S., and Noegel, A.A. (1999) RacF1, a novel member of the Rho protein family in Dictyostelium discoideum, associates transiently with cell contact areas, macropinosomes, and phagosomes. Mol. Biol. Cell 10, 1205–1219.
Seastone, D.J., Lee, E., Bush, J., Knecht, D., and Cardelli, J. (1998) Overexpression of a novel Rho family GTPase, RacC, induces unusual actin-based structures and positively affects phagocytosis in Dictyostelium discoideum. Mol. Biol. Cell 9, 2891–2904.
Hall, A. L., Schlein, A., and Condeelis, J. (1988) Relationship of pseudopod extension to chemotactic hormone-induced actin polymerization in amoeboid cells. J. Cell. Biochem. 37, 285–299.
Howard, T.H., and Oresajo, C.O. (1985) The kinetics of chemotactic peptide-induced change in F-actin content, F-actin distribution, and the shape of neutrophils. J Cell Biol 101, 1078–1085.
Benard, V., Bohl, B.P., and Bokoch, G.M. (1999) Characterization of rac and cdc42 activation in chemoattractant-stimulated human neutrophils using a novel assay for active GTPases. J Biol Chem 274, 13198–13204.
Vlahou, G., Schmidt, O., Wagner, B., Uenlue, H., Dersch, P., Rivero, F., and Weissenmayer, B.A. (2009) Yersinia outer protein YopE affects the actin cytoskeleton in Dictyostelium discoideum through targeting of multiple Rho family GTPases. BMC Microbiol 9, 138.
de la Roche, M., Mahasneh, A., Lee, S.F., Rivero, F., and Cote, G.P. (2005) Cellular distribution and functions of wild-type and constitutively activated Dictyostelium PakB. Mol Biol Cell 16, 238–247.
Condeelis, J., and Hall, A.L. (1991) Measurement of actin polymerization and cross-linking in agonist-stimulated cells. Methods Enzymol 196, 486–496.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Xiong, H., Rivero, F. (2012). Assaying Rho GTPase-Dependent Processes in Dictyostelium discoideum . In: Rivero, F. (eds) Rho GTPases. Methods in Molecular Biology, vol 827. Springer, New York, NY. https://doi.org/10.1007/978-1-61779-442-1_25
Download citation
DOI: https://doi.org/10.1007/978-1-61779-442-1_25
Published:
Publisher Name: Springer, New York, NY
Print ISBN: 978-1-61779-441-4
Online ISBN: 978-1-61779-442-1
eBook Packages: Springer Protocols