Two regions with differential growth-modulating activity in the N-terminal domain of ras GTPase-activating protein (p120GAP) src homology and Gly-Ala-Pro-rich regions.
Ras GTPase-activating protein of 120 kDa (p120GAP) consists of a hydrophobic Gly-Ala-Pro-rich stretch and src homology 2 and 3 (SH2/SH3) domains in the N-terminal half, and a Ras GTPase-activating domain at the C-terminus. In order to evaluate the potential for cell-growth regulation of the N-terminal region of p120GAP, we isolated three distinct clones of rat 3Y1 fibroblast that express either the SH2/SH3 regions alone, the N-terminal half, or the whole p120GAP. Clones that express the SH2-SH3-SH2 regions of 37 kDa (p37SH2/3) at a level of only 15-30% that of endogenous p120GAP, but not clones expressing complete p120GAP or its N-terminal half of 55 kDa (p55GAP-N), showed significant growth-enhancing properties, including a higher saturation density and increased uptake of 2-deoxyglucose. Clones expressing p37SH2/3 or p55GAP-N maintained high levels of tyrosine-phosphorylated p190 and p62, both of which bind the SH2 domain of p120GAP, while clones expressing the whole p120GAP showed no tyrosine phosphorylation of p62. Furthermore, in the presence of a phorbol ester, only the clones expressing p37SH2/3 showed increased tyrosine phosphorylation of p62 and c-fos expression. These clones also showed the ability of colony formation in soft agar. These results indicate that the N-terminal domain of p120GAP consists of two regions with differential growth-enhancing activities and suggest that the transforming potential of SH2/SH3 regions is blocked by the N-terminal hydrophobic Gly-Ala-Pro stretch.
Chang
J S
JS
Mitsubishi Kasei Institute of Life Sciences, Tokyo, Japan.
Kobayashi
M
M
Wang
D Z
DZ
Maruta
H
H
Iwashita
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002455:Cell Division; D002471:Cell Transformation, Neoplastic / Q000737:chemistry; D002999:Clone Cells; D003847:Deoxyglucose / Q000493:pharmacokinetics; D020690:GTPase-Activating Proteins; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011493:Protein Kinase C / Q000502:physiology; D011506:Proteins / Q000737:chemistry* / Q000502:physiology; D051381:Rats; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D020703:ras GTPase-Activating Proteins; D018909:src Homology Domains
D020690:GTPase-Activating Proteins; D011506:Proteins; D020703:ras GTPase-Activating Proteins; D014443:Tyrosine; D003847:Deoxyglucose; D011493:Protein Kinase C