Figure 3

Endogenous EGFR in NSCLC cells induces tyrosine phosphorylation on GPRC5A. A, Western blot of eleven NSCLC cell lines was performed with anti-GPRC5A, anti-EGFR, anti-HER-2, and anti-actin, as indicated. B, H292G Cells were serum-starved for 24 hours, then treated with or without EGF (100 ng/ml) for 5 min. Cell lysates were harvested for IP-Western using either anti-GPRC5A or normal IgG. Western blot was performed using either PY99 antibody or anti-GPRC5A as indicated. C. H1792-GPRC5A, D, H1792-V, H1792-5A, H1792-4 F stable transfectants were serum-starved for 24 hours and then treated with EGF for 5 minutes or pretreated with AG1478 (30 ng/ml) for 6 hours as indicated. Total cellular proteins were analyzed by IP-Western using the antibodies as indicated.