Fig. 4.

EGR1 induction by ER stress is dependent on RAS, RAF1, and SRC. HepG2 cells were transiently transfected with 100 nM siRNA, as described in the Methods section, and cultured for 48 h. Cells were incubated in DMEM ± Tg for 6 h and then steady state mRNA for the indicated gene was measured by RT-qPCR. (Panel A) Cells were transfected with an siRNA against a non-targeting control (Ctrl), or H, K, or N RAS, and then mRNA was measured for EGR1, each RAS form, and GAPDH as the internal control. (Panel B) HepG2 cells were transfected with an siRNA against a non-targeting control (Ctrl) or against RAF1 and then mRNA was measured for EGR1, RAF1, and GAPDH as the internal control. (Panel C) Cells were transfected with an siRNA against a non-targeting control (Ctrl) or siRNA against SRC and then mRNA was measured for EGR1, SRC, and GAPDH as the internal control. For all panels, the target mRNA values were normalized to those for GAPDH within the same sample and the results shown are the means ± SD of triplicate samples within an experiment. The results shown are representative of multiple experiments. The data are graphed as relative values setting the siControl DMEM value to 1.0. An asterisk indicates that the targeted siRNA value is significantly different (p ≤ 0.05) from the corresponding siControl.