Stau1 levels regulate the pre-mRNA splicing of the human <i>INSR</i> in HeLa cells.

<p>(A) pGIPZ (CTRL) or Stau1-HA (Stau1-HA) plasmids were transiently transfected into HeLa cell lines, and total RNA and protein lysate was collected after 48 hours. RT-PCR using primers specific to the human endogenous <i>INSR</i> were used on cDNA synthesized from total RNA to amplify the two isoforms (IR-A and IR-B) of the <i>INSR</i>. Stau1-HA protein levels were assessed by Western blot using HA-specific antibodies, and GAPDH was used as a loading control. (B) shCTRL or shStau1 were transiently transfected into HeLa cell lines and total RNA and protein lysate was collected after 48 hours. RT-PCR was performed to amplify the <i>INSR</i> isoforms. Stau1 protein levels were assessed by Western blot and quantified using GAPDH as a loading control. (C) Representative Western blots showing protein levels of CUGBP1, MBNL1 and hnRNP H in HeLa cells transfected with CTRL, shRNA or Stau1-HA plasmids. GAPDH was used as a loading control. (D) Quantification of Western blot analysis of splicing factors upon Stau1 level modulation. In all cases, bar graphs show an average of ≥3 independent experiments. Error bars represent SEM * = p < 0.05, ** = p < 0.01.