RPS12 overexpression affects heavy polysome association of APAF-1 and XIAP mRNAs. (A) Semi-quantitative RT-PCR was used to measure the abundance of APAF-1, XIAP, GAPDH, and β-Actin mRNAs in polysome fractions of normoxic (21% O2) and hypoxic (1% O2) HEK293 cells overexpressing RPS12 (eS12) with the Myc-DDK-RPS12 ORF vector (red line) or control cells expressing the empty vector pMyc-N1 (blue line). The fraction of mRNA in each lane was calculated based on total band intensity of the transcript across the entire polysome gradient. Dashed line denotes the start of the polysome fractions. A representative image is shown. (B) Total levels of APAF-1 and XIAP mRNA were measured in normoxic and hypoxic HEK293 control cells and those overexpressing RPS12 using RT-qPCR. The ΔΔCt method was used, normalizing to reference gene RPL13A, and fold-change made relative to empty vector control. Data (n = 3), mean ± SEM. Two-tailed unpaired t-tests were performed.
