Identification of sites on epidermal growth factor receptors which are phosphorylated by pp60src in vitro.
The Epidermal Growth Factor Receptor (EGF-R) becomes constitutively tyrosine phosphorylated on two novel sites in v-Src transformed cells, and these phosphorylations are associated with enhanced signaling activity [1]. To determine whether Src could directly phosphorylate these sites, we have examined the ability of the Src kinase to phosphorylate both wild-type and kinase-defective EGF-Rs in vitro. Although purified Src could phosphorylate EGF-Rs, the pattern of phosphorylation sites was not identical to what was previously found in vivo [1]: Src in vitro directly phosphorylated EGF-Rs on one autophosphorylation site (Tyr 1173) which was not a site of re-induced in vivo phosphorylation, suggesting the in vivo inaccessibility of this site. One Src-specific in vitro phosphorylation site (Tyr 03) appeared to correspond to one of the in vivo Src-induced sites (sPY2), but the other Src-specific in vivo site (sPY1) was not significantly phosphorylated in vitro, raising the possibility of a Src-induced tyrosine kinase cascade. The ability of Src to phosphorylate the EGF-R is consistent with the suggestion that the receptor can function as a kinase substrate independent of its intrinsic enzymatic activity, as implied by recent studies on signaling by kinase-defective EGF-Rs.
Wright
J D
JD
Department of Pharmacology, University of Virginia Health Sciences Center, Charlottesville 22908, USA.
Reuter
C W
CW
Weber
M J
MJ
D000595:Amino Acid Sequence; D001665:Binding Sites; D002460:Cell Line; D002461:Cell Line, Transformed; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004591:Electrophoresis, Polyacrylamide Gel; D066246:ErbB Receptors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D015688:Oncogene Protein pp60(v-src) / Q000378:metabolism / Q000494:pharmacology; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D010455:Peptides; D014443:Tyrosine; D066246:ErbB Receptors; D020794:Receptor Protein-Tyrosine Kinases; D015688:Oncogene Protein pp60(v-src); D016392:Proto-Oncogene Proteins pp60(c-src); D014357:Trypsin