TYROSINE PHOSPHORYLATION OF RAS GTPASE-ACTIVATING PROTEIN DOES NOT REQUIRE ASSOCIATION WITH THE EPIDERMAL GROWTH-FACTOR RECEPTOR

Citation
C. Soler et al., TYROSINE PHOSPHORYLATION OF RAS GTPASE-ACTIVATING PROTEIN DOES NOT REQUIRE ASSOCIATION WITH THE EPIDERMAL GROWTH-FACTOR RECEPTOR, The Journal of biological chemistry, 268(29), 1993, pp. 22010-22019
Citations number
71
Categorie Soggetti
Biology
ISSN journal
00219258
Volume
268
Issue
29
Year of publication
1993
Pages
22010 - 22019
Database
ISI
SICI code
0021-9258(1993)268:29<22010:TPORGP>2.0.ZU;2-5
Abstract
The importance of the carboxyl-terminal domain of the epidermal growth factor (EGF) receptor and its five autophosphorylation sites in the i n vivo interaction and tyrosine phosphorylation of the ras GTPase-acti vating protein (rasGAP) has been investigated, using NIH 3T3 cells tra nsfected with mutant EGF receptors. Phosphorylation of rasGAP by EGF r eceptor mutants, in which one to four autophosphorylation sites (Tyr-1 173, -1148, -1086, and -1068) were mutated to phenylalanine, was reduc ed by 50-60% compared to the wild-type receptor. Elimination of these four autophosphorylation sites by truncation of 123 carboxyl-terminal residues of the EGF receptor paralleled results obtained with point mu tants. Substantial inhibition (about 90%) of rasGAP tyrosine phosphory lation by the EGF receptor occurred only when the remaining autophosph orylation site (Tyr-1992) was mutated, in the context of this truncate d receptor or in the full-length receptor mutated at all four other au tophosphorylation sites. However, a point mutation of only Tyr-992 in the full-length receptor suppressed tyrosine phosphorylation of rasGAP only by 50%. In contrast, an EGF receptor lacking the last 214 amino acid residues (Dc214), which emcompasses all five autophosphorylation sites, phosphorylated rasGAP to the same extent as the wild-type recep tor. However, this truncated receptor was significantly impaired in it s capacity to phosphorylate phospholipase C-gamma1. Interestingly, whi le EGF receptor autophosphorylation sites are required for EGF-induced rasGAP association with the receptor, maximal phosphorylation of rasG AP by the truncated receptor Dc214 occurred without detectable formati on of receptor.rasGAP complexes. Furthermore, the capacity of mutated EGF receptors to bring about focal transformation was correlated with their capacity to phosphorylate rasGAP.