Identification of the SH3 domain of GAP as an essential sequence for Ras-GAP-mediated signaling.
Guanosine triphosphatase activating protein (GAP) is an essential component of Ras signaling pathways. GAP functions in different cell types as a deactivator and a transmitter of cellular Ras signals. A domain (amino acids 275 to 351) encompassing the Src homology region 3 (SH3) of GAP was found to be essential for GAP signaling. A monoclonal antibody was used to block germinal vesicle breakdown (GVBD) induced by the oncogenic protein Ha-ras Lys12 in Xenopus oocytes. The monoclonal antibody, which was found to recognize the peptide containing amino acids 275 to 351 within the amino-terminal domain of GAP, did not modify the stimulation of the Ha-Ras-GTPase by GAP. Injection of peptides corresponding to amino acids 275 to 351 and 317 to 326 blocked GVBD induced by insulin or by Ha-Ras Lys12 but not that induced by progesterone. These findings confirm that GAP is an effector for Ras in Xenopus oocytes and that the SH3 domain is essential for signal transduction.
Duchesne
M
M
Rhone Poulenc Rorer, Centre de Recherche de Vitry-Alfortville, Vitry Sur Seine, France.
Schweighoffer
F
F
Parker
F
F
Clerc
F
F
Frobert
Y
Y
Thang
M N
MN
Tocqué
B
B
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D003001:Cloning, Molecular; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000378:metabolism; D020690:GTPase-Activating Proteins; D011905:Genes, ras; D016391:Genes, src; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D009865:Oocytes / Q000502:physiology; D016133:Polymerase Chain Reaction / Q000379:methods; D011506:Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014981:Xenopus; D020703:ras GTPase-Activating Proteins
D000911:Antibodies, Monoclonal; D000939:Epitopes; D020690:GTPase-Activating Proteins; D011506:Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D020703:ras GTPase-Activating Proteins; D005982:Glutathione Transferase; D020558:GTP Phosphohydrolases