Molecular analysis of Ras activation by tyrosine phosphorylated Vav.
Vav has been shown to activate Ras (1-3) and is regulated by tyrosine phosphorylation (1) or binding of diglycerides (3) to the cysteine rich domain. In the present study employing different Ras activation assay techniques using [3H]GDP release or [32P]alpha GTP-binding from membrane-bound or soluble recombinant Ras, we demonstrate that Ras activity can be increased by tyrosine phosphorylated Vav upon cellular stimulation via the IL-2 receptor or the TCR/CD3-complex. Increase of [32P]alpha GTP-binding to Ras catalyzed by phosphorylated Vav is similar to the activity of immunoprecipitated Sos. The activity of Vav measured by binding of [32P]alpha GTP to Ras was linear with respect to the concentration of Vav protein used. To study molecular characteristics of this Vav-Ras interaction, we used several Ras mutants and demonstrate that Vav activity towards Ras depends on the integrity of the same or similar domains as Ras activation by SDC 25 or CDC 25.
Gulbins
E
E
I. Institute of Physiology, University of Tuebingen, Germany.
Schlottmann
K
K
Brenner
B
B
Lang
F
F
Coggeshall
K M
KM
D017252:CD3 Complex / Q000378:metabolism; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051567:Proto-Oncogene Proteins c-vav; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D015375:Receptors, Interleukin-2 / Q000378:metabolism; D015398:Signal Transduction
D017252:CD3 Complex; D018797:Cell Cycle Proteins; D011518:Proto-Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; D015375:Receptors, Interleukin-2; C496618:VAV1 protein, human; D019000:Phosphotyrosine; D006160:Guanosine Triphosphate; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)