Post-transcriptional regulation of LUC7L3 by TRAP1. (A) Assessment of LUC7L3 levels upon TRAP1 modulation. (Left) WB analysis of cell extracts upon 72 h of silencing using TRAP1-directed siRNA (PEA1 and HCT116) or TRAP1-directed shRNA induction using tetracycline (HeLa FITR). (Right) Relative densitometric analysis calculated by assuming protein levels of the control equal to 1. Statistically significant differences were detected using two-tailed paired Student's t-test (PEA1 n = 5, HCT116 n = 3, HeLa n = 3). Statistical significance is represented as follows: (*) P < 0.05, (**) P < 0.01, and (***) P < 0.001. Error bars represent SEM. (B) Polysome profiling absorbance, measured at 254 nm, of PEA1 cell extracts, from control and siTRAP1 cells. (C) Quantification of the polysome/monosome area ratio in control and siTRAP1 profiles as reported in A (n = 4). Statistically significant differences were detected using two-tailed unpaired Student's t-test. Error bars represent SEM. (D) Percentages of indicated mRNA distributed in sucrose gradient fractions of control and siTRAP1 cells (n = 3). Data are reported as mean ± SEM. Individual P-values are indicated according to two-way ANOVA test. (E) LUC7L3 mRNA levels do not decrease after TRAP1 depletion. Transcript levels were assessed by qRT-PCR, corrected for PPIA, and normalized to the levels in shCTRL cells (red line; n = 3). Significance was assessed by one-sample t-test. Statistical significance is represented as follows: (*) P < 0.05, (**) P < 0.01, (***) P < 0.001. Error bars represent SEM.
