W. Lopaczynski et C. Terry, Insulin-like growth factor I activates insulin receptor substrate 1 and Ras in human osteosarcoma cells, ACT BIOCH P, 46(1), 1999, pp. 117-123
Insulin-like growth factor I (IGF-I) stimulates multiplication of the human
osteosarcoma cell line, MG-63, by acting through IGF-I receptor. We have c
haracterized IGF-I stimulated phosphorylation of IRS-1, activation of Ras c
ycle and phosphorylation of c-Jun in this cell line. Serum starved MG-63 ce
lls were (1) IGF-I stimulated and lysates were immunoprecipitated with poly
clonal IRS-1 antibody or (2) metabolically labeled with [P-32]orthophosphor
ic acid and then cells were treated with IGF-I. Cell lysates were immunopre
cipitated with p21Ras antibody (Y13-259) and bound nucleotides were analyse
d by thin-layer chromatography. We demonstrated tyrosine phosphorylation of
IRS-1/2 immunoprecipitated from MG-63 cells stimulated with IGF-I. We also
showed an increased level of GTP in p21Ras immunoprecipitates from IGF-I t
reated cells. Nuclear extracts prepared from P-32-labeled cells before and
after addition of IGF-I were immunoprecipitated with c-Jun antibody. After
electrophoresis and autoradiography, phosphorylation of the c-Jun band was
seen to be IGF-I independent. Phosphoamino acid analysis of the c-Jun band
showed that phosphoserine was the major species.