在盘基网柄菌中检测Rho GTP酶依赖性过程。

Assaying Rho GTPase-Dependent Processes in Dictyostelium discoideum.

作者信息

Marinović Maja, Xiong Huajiang, Rivero Francisco, Weber Igor

机构信息

Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia.

Department of Biochemistry, University of Oxford, Oxford, UK.

出版信息

Methods Mol Biol. 2018;1821:371-392. doi: 10.1007/978-1-4939-8612-5_25.

DOI:10.1007/978-1-4939-8612-5_25

PMID:30062425

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 cells make use of Rho-regulated signaling pathways to reorganize the actin 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, and for monitoring the localization and dynamics of Rac activity in live cells.

摘要

模式生物盘基网柄菌非常适合用于研究分子和细胞生物学的基本问题，尤其是那些与细胞骨架的结构、调控和动力学、信号转导、细胞间黏附以及发育相关的问题。盘基网柄菌细胞在趋化作用、内吞作用和胞质分裂过程中利用Rho调节的信号通路来重组肌动蛋白细胞骨架。在这种生物中，Rho家族包含20个成员，其中几个属于Rac亚家族，但没有Cdc42和Rho亚家族的代表。在这里，我们提供了适用于监测有聚集能力的细胞在用趋化剂cAMP刺激时肌动蛋白聚合反应和Rac激活的方案，以及用于监测活细胞中Rac活性的定位和动力学的方案。