Ect2 and MgcRacGAP regulate the activation and function of Cdc42 in mitosis

J Cell Biol. 2005 Jan 17;168(2):221-32. doi: 10.1083/jcb.200408085. Epub 2005 Jan 10.

Abstract

Although Rho regulates cytokinesis, little was known about the functions in mitosis of Cdc42 and Rac. We recently suggested that Cdc42 works in metaphase by regulating bi-orient attachment of spindle microtubules to kinetochores. We now confirm the role of Cdc42 by RNA interference and identify the mechanisms for activation and down-regulation of Cdc42. Using a pull-down assay, we found that the level of GTP-Cdc42 elevates in metaphase, whereas the level of GTP-Rac does not change significantly in mitosis. Overexpression of dominant-negative mutants of Ect2 and MgcRacGAP, a Rho GTPase guanine nucleotide exchange factor and GTPase activating protein, respectively, or depletion of Ect2 by RNA interference suppresses this change of GTP-Cdc42 in mitosis. Depletion of Ect2 also impairs microtubule attachment to kinetochores and causes prometaphase delay and abnormal chromosomal segregation, as does depletion of Cdc42 or expression of the Ect2 and MgcRacGAP mutants. These results suggest that Ect2 and MgcRacGAP regulate the activation and function of Cdc42 in mitosis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autoantigens / metabolism
  • Calcium-Binding Proteins / metabolism
  • Cell Cycle / drug effects
  • Cell Cycle / physiology
  • Cell Cycle Proteins
  • Centromere Protein A
  • Chromosomal Proteins, Non-Histone / metabolism
  • GTPase-Activating Proteins / genetics
  • GTPase-Activating Proteins / metabolism
  • GTPase-Activating Proteins / physiology*
  • HeLa Cells
  • Humans
  • Kinetics
  • Kinetochores / metabolism
  • Mad2 Proteins
  • Metaphase / physiology
  • Microscopy, Fluorescence
  • Microtubule-Associated Proteins / metabolism
  • Mitosis / physiology*
  • Mutation
  • Nocodazole / pharmacology
  • Prometaphase / physiology
  • Protamine Kinase / metabolism
  • Proto-Oncogene Proteins / genetics
  • Proto-Oncogene Proteins / metabolism
  • Proto-Oncogene Proteins / physiology*
  • RNA Interference / physiology
  • RNA, Double-Stranded / genetics
  • Repressor Proteins
  • Spindle Apparatus / metabolism
  • Thymidine / pharmacology
  • Transfection
  • Tubulin / metabolism
  • cdc42 GTP-Binding Protein / metabolism
  • cdc42 GTP-Binding Protein / physiology*
  • rac GTP-Binding Proteins / metabolism

Substances

  • Autoantigens
  • Calcium-Binding Proteins
  • Cell Cycle Proteins
  • Centromere Protein A
  • Chromosomal Proteins, Non-Histone
  • EB1 microtubule binding proteins
  • ECT2 protein, human
  • GTPase-Activating Proteins
  • MAD2L1 protein, human
  • Mad2 Proteins
  • Microtubule-Associated Proteins
  • Proto-Oncogene Proteins
  • RNA, Double-Stranded
  • Repressor Proteins
  • Tubulin
  • mgcRacGAP
  • Protamine Kinase
  • histone h2b kinase
  • cdc42 GTP-Binding Protein
  • rac GTP-Binding Proteins
  • Nocodazole
  • Thymidine