Characterization of the kinase activity essential for tyrosine phosphorylation of p130Cas in fibroblasts.
The cellular transformation by v-Src or v-Crk induces tyrosine phosphorylation of a common substrate molecule, p130Cas (Cas), which tightly binds these oncoproteins in vivo. From its structure, Cas is deduced to be an ideal substrate for Src family kinases and Abl kinase. The tyrosine kinase activity associated with Cas was analysed using mouse variant fibroblasts lacking at least one of tyrosine kinases. In normal fibroblasts, Cas is associated with a significant level of tyrosine kinase activity which efficiently phosphorylates Cas in vitro. The Cas-associated tyrosine kinase activity was remarkably elevated in Csk-/- cells, which resulted in hyperphosphorylation of cellular Cas. The associated kinase activity was slightly increased in Src-/- cells whereas not significantly changed in Abl-/- nor Fak-/- cells. On the contrary, the Cas-associated kinase activity was remarkably decreased in Fyn-/- cells. At the same time, association of Cas with Fyn kinase in vitro was most obviously detected in normal fibroblasts as well as Csk-/- cells. Transient expression of v-Crk induced elevation of the Cas-associated kinase activity in all of these cell lines except the primary culture of Fyn-/- fibroblasts. These results indicate that Fyn kinase plays an essential role in v-Crk-mediated phosphorylation of Cas.
Sakai
R
R
Molecular Biology Division, Jichi Medical School, Kawachi-gun, Tochigi, Japan.
Nakamoto
T
T
Ozawa
K
K
Aizawa
S
S
Hirai
H
H
D016475:3T3 Cells; D000818:Animals; D002478:Cells, Cultured; D029563:Cellular Apoptosis Susceptibility Protein; D051379:Mice; D050736:Oncogene Protein v-crk; D010449:Peptide Mapping; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D015682:Retroviridae Proteins, Oncogenic / Q000378:metabolism*; D018909:src Homology Domains
D029563:Cellular Apoptosis Susceptibility Protein; D050736:Oncogene Protein v-crk; D011506:Proteins; D015682:Retroviridae Proteins, Oncogenic; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases