Membrane-targeting is critical for the phosphorylation of Vav2 by activated EGF receptor.
Vav2 is a member of the Vav family that serves as guanine nucleotide exchange factors (GEFs) for the Rho family of Ras-related GTPases. Unlike Vav1, whose expression is restricted to cells of hematopoietic origin, Vav2 is broadly expressed. Recently, Vav2 has been identified as a substrate for the EGF receptor. Here, we show that in EGF-treated COS7 cells Vav2 is phosphorylated on tyrosine residues and associates with the EGF receptor. In addition, introducing point mutations into the SH2 domain of green fluorescens protein (GFP)-Vav2 fusion protein leads to the loss of Vav2 tyrosine phosphorylation in response to EGF. To investigate further the mechanism of Vav2 phosphorylation, N-terminal (NT) domain of Vav2 was transiently expressed in COS7 cells as GFP fusion protein. Whereas the NT domain of Vav2 is a preferred substrate for the activated EGF receptor in vitro, we could not detect tyrosine phosphorylation of the GFP-NT construct in EGF-treated cells. However, when the SH2 domain of Vav2 was fused to its NT domain, NT domain proved to be a substrate for the EGF receptor in vivo. These data suggest that membrane-targeting of Vav2 through its SH2 domain is an important event in the phosphorylation and activation of Vav2 in response to EGF.
Tamás
P
P
Department of Medical Chemistry, Semmelweis University Medical School, 9 Puskin Street, 1088, Budapest, Hungary.
Solti
Z
Z
Buday
L
L
D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D009154:Mutation; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction
D008164:Luminescent Proteins; D015513:Oncogene Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D019000:Phosphotyrosine; D066246:ErbB Receptors