Figure 8. Effects of overexpression of CFIm59 on matrix stiffness–regulated APA and expression of COL1A1 and FN1.

(A) Human lung fibroblasts were infected by recombinant lentiviral vectors expressing CFIm59-GFP or GFP alone. Ectopic expression of CFIm59-GFP fusion protein and GFP protein was determined by immunoblot with both anti-CFIm59 and anti-GFP antibodies. GAPDH was used as loading control. (B–G) Lung fibroblasts infected by CFIm59-GFP or GFP viruses were cultured on soft and stiff hydrogels for 48 hours. Ratios of long-to-total COL1A1 (B) and FN1 (E) transcripts were used to evaluate the proximal poly(A) site usage. Relative levels of COL1A1 (C) and FN1 (F) mRNA were determined by real-time RT-PCR. Protein levels of COL1A1 (D) and FN1 (G) were determined by immunoblot. GAPDH was used as internal reference or loading control. Bar graphs represent (mean ± SD) 3 or 4 separate experiments. Statistical analysis was performed by 1-way ANOVA.