Constitutive phosphorylation of 70z-Cbl when
expressed in MDCK cells. (A) Scheme of 70z-Cbl and its mutant variants.
PTB/SH2, phosphotyrosine-binding SH2 domain; RF, RING finger domain;
Pro, proline-rich domain. (B) 70z-Cbl and its variants were
immunoprecipitated from MDCK cell populations, separated on an 8%
SDS-PAGE gel, and immunoblotted with anti-HA (upper panel)
or anti-phosphotyrosine (lower panel) antibodies. (C) Lysates from MDCK
cell populations were incubated with the amino-terminal SH2 domain of
p85 (GST-N-SH2 p85; upper panel) or the amino- and carboxyl-terminal
SH3 domains of Grb2 (GST-N+C-SH3 Grb2; lower panel). Proteins were
separated on an 8% SDS-PAGE gel, transferred to nitrocellulose, and
immunoblotted with anti-HA antibody. (D) MDCK cell
populations were immunoprecipitated with the indicated antibodies,
resolved by SDS-PAGE, transferred to nitrocellulose, and
immunoblotted with anti-HA antibody.