Figure 1

FASN phosphorylation by HRG in SKBR3 cells. (A) SKBR3 cells were incubated in serum-free medium overnight and then treated with 50 ng/mL heregulin (HRG) for the indicated times. FASN was immunoprecipitated and immunoblotted by the use of a monoclonal phosphotyrosine (PT66) and anti-FASN antibodies. (B) Immunoprecipitated FASN substrate was incubated with HER2 kinase and radiolabeled with (γ-P³²)-ATP for 30 minutes at 30ºC, separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and visualized by autoradiography. Recombinant HER2 kinase was used as a positive control. Western blotting for FASN and HER2 was performed to confirm that equal amounts of FASN or HER2 were used in each kinase assay.