Ezrin function is required for ROCK-mediated fibroblast transformation by the Net and Dbl oncogenes.
The small G protein RhoA and its GDP/GTP exchange factors (GEFs) Net and Dbl can transform NIH 3T3 fibroblasts, dependent on the activity of the RhoA effector kinase ROCK. We investigated the role of the cytoskeletal linker protein ezrin in this process. RhoA effector loop mutants which can bind ROCK induce relocalization of ezrin to dorsal actin-containing cell surface protrusions, as do Net and Dbl. Both processes are inhibited by the ROCK inhibitor Y27632, which also inhibits association of ezrin with the cytoskeleton, and phosphorylation of T567, conserved between ezrin and its relatives radixin and moesin. ROCK can phosphorylate the ezrin C-terminus in vitro. The ezrin mutant T567A cannot be relocalized by activated RhoA, Net or Dbl or by ROCK itself, and also inhibits RhoA-mediated contractility and focal adhesion formation. Moreover, ezrin T567A, but not wild-type ezrin, restores contact inhibition to Net- and Dbl-transformed cells, and inhibits the activity of Net and Ras in focus formation assays. These results implicate ROCK-mediated ezrin C-terminal phosphorylation in transformation by RhoGEFs.
Tran Quang
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Transcription Laboratory, Room 528, Imperial Cancer Research Fund Laboratories, 44 Lincoln's Inn Fields, London WC2A 3PX, UK.
Gautreau
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Arpin
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Treisman
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D016475:3T3 Cells; D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D003598:Cytoskeletal Proteins; D003599:Cytoskeleton / Q000201:enzymology; D020662:Guanine Nucleotide Exchange Factors; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D015513:Oncogene Proteins; D009857:Oncogenes; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D050783:Proto-Oncogene Proteins c-ets; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics; D015398:Signal Transduction; D014157:Transcription Factors / Q000235:genetics; D054460:rho-Associated Kinases; D000099149:Ezrin
D003598:Cytoskeletal Proteins; D020662:Guanine Nucleotide Exchange Factors; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; D010750:Phosphoproteins; D017346:Protein Serine-Threonine Kinases; D050783:Proto-Oncogene Proteins c-ets; D015682:Retroviridae Proteins, Oncogenic; D014157:Transcription Factors; D054460:rho-Associated Kinases; D000099149:Ezrin; C086924:Elk3 protein, mouse; C512843:Mcf2 protein, mouse