FIG. 8.
Ectopic expression of PKCδ affects SRE-dependent transcription and endogenous immediate-early gene expression. (A) A PKCδ Western blot (sc-937) with extracts of young and senescent cells previously transfected with the pkV vector (V), PKCδ-FL (FL), PKCδ-CF (CF), or dominant negative PKCδ-CF(K-R) (DN). (B) Ectopic expression of PKCδ constructs and RT-PCR analysis of endogenous c-fos transcript levels in young and senescent fibroblast RNA. Glyceraldehhyde-3-phosphate dehydrogenase (GAPDH) served as an internal control for loading, amplification, efficiency, and RNA integrity. (C and D) Ectopic expression of PKCδ constructs and SRE-driven reporter expression in young (C) and senescent (D) fibroblasts. Young and senescent fibroblasts were transfected with equal amounts of a c-fos luciferase reporter and PKCδ expression constructs. Luciferase activity was measured before and after serum induction for each cotransfection to assess c-fos SRE- plus ets-, SRE-, or ets-dependent transcription in vivo. The histograms show the effect of induction relative to each serum-starved transfected construct for three independent trials. Error bars indicate standard deviations.