Figure 6.

cAMP-Stimulated Expression of Genes Involved in Cholesterol Synthesis and Uptake Is Not Mediated by Accumulation of Processed SREBP2

A, Relative mRNA levels in MA-10 cells treated with combinations of 50 μm 8-Br-cAMP and 10 μm 25-OHC. B, These concentrations of 25-OHC are capable of blocking cholesterol-starved HEK-293 cells from activating SREBP2. The 125-kDa precursor form (P) of SREBP2 and 70-kDa nuclear form (N) are visible in whole-cell extracts of lipid-starved HEK-293 cells. These data are representative of duplicate experiments. C, Relative mRNA levels in MA-10 cells treated with combinations of 50 μm 8-Br-cAMP and 5 μg/ml AG. Each bar represents the mean expression level for three independent experiments. Letters indicate P < 0.05 for a one-way ANOVA followed by Newman-Keuls post test. Levels of mRNA were normalized between samples based on the levels of Actb transcript. D, Treatment of MA-10 cells with cAMP or DHT, alone or in combination, has no effect on levels of the 68-kDA nuclear form of SREBP2. These data are representative of experiments performed in triplicate. E, SREBP2 is not processed in Leydig cell-enriched protein extracts from wild-type or Ar^{invflox(ex1-neo)/Y} testes. F, HMGCR levels are elevated in Leydig cell-enriched protein extracts from wild-type or Ar^{invflox(ex1-neo)/Y} testes. Cells used to generate protein extracts in E and F contained more than 60% Leydig cells as determined by 3βHSD cytochemistry.