FIG. 1.

Knockdown of RalGDS suppresses activation of Akt by growth factors in MCF10A cells. (A) Expression levels of RalGDS RNA in MCF10A cells stably expressing RalGDS shRNA or empty vector detected by RT-PCR. (B) MCF10A cells stably expressing either RalGDS shRNA or empty vector were serum starved for 6 h and then stimulated with insulin (1 μM) for the indicated times, and cell lysates were assayed for pAkt308, pAkt473, pERK, and total Akt. pAkt308 band intensities were quantified with NIH ImageJ 1.34s, and the data represent the averages of data from three independent experiments (+ standard deviations). (C) MCF10A cells were stimulated with EGF (5 ng/ml) after being starved for 6 h and then analyzed as in panel B.