Skip to main content
. 2010 Dec 1;286(6):4173–4185. doi: 10.1074/jbc.M110.157420

FIGURE 4.

FIGURE 4.

Knock down of PTP1B expression by siRNA results in an increase in the level of tyrosine phosphorylation of PLC-γ1, Gab1, and SHP2. HEK293 stably expressing PTP1B siRNA (lanes 3 and 4) or a scrambled control (lanes 1 and 2) were transiently transfected with a plasmid encoding the constitutively active Src Y527F (lanes 2 and 4) or with the empty vector (lanes 1 and 3). A, protein extracts were immunoblotted with the 4G10 antibody, to monitor the phosphotyrosine profile of HEK293 cells, with an antibody against the Src kinase to verify ectopic expression of the constitutively active mutant (lanes 2 and 4), with an anti-PTP1B antibody, to monitor the endogenous PTP1B expression levels and antitubulin, as a loading control. B, 1 mg of protein extract was immunoprecipitated with the anti-phosphotyrosine antibody 4G10 (IP: anti-4G10). The input cell extracts and the IP material were immunoblotted (WB) with antibodies against PLC-γ1, Gab1, SHP2, and Grb2. The amount of IgG was used as loading control. The phosphorylation levels of the indicated proteins were normalized and plotted as a bar diagram.