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. Author manuscript; available in PMC: 2017 Jun 26.
Published in final edited form as: Cell Rep. 2016 Nov 15;17(8):1990–2003. doi: 10.1016/j.celrep.2016.10.071

Figure 2. SRSF10 Requires hnRNP F/H to Control Bcl-x Splicing and Interacts with hnRNP F, H, and K Proteins.

Figure 2

(A) Schematic representation of regulatory elements surrounding the 5′ splice site of Bcl-xS. (B) The X2 minigene and derivatives were transfected either alone or with the HA-SRSF10 plasmid. RT-PCR assays were carried out, and radiolabeled RT-PCR products were fractionated on a non-denaturing gel to quantitate Bcl-xS and Bcl-xL products. The histograms represent the average percentage of Bcl-xS products from triplicate experiments. (C) siF/H was used to deplete hnRNP F/H with an immunoblot shown to verify depletion. HA-SRSF10 was transfected in 293 cells treated or not with siF/H. RT-PCR was carried out to detect the Bcl-x splice variants, as described in (B). (D) Immunoprecipitation assays using 293 cells transfected with FLAG-SRSF10. The material recovered was fractionated and transferred on nitrocellulose decorated with anti-FLAG antibodies. (E) siK was used to deplete hnRNP K and an immunoblot is shown. HA-SRSF10 was transfected in 293 cells treated or not with siK. RT-PCR was carried out to detect the Bcl-x splice products, as described in (B). (F) Immunoprecipitation assays with hnRNP K antibody. Using 293 cells transfected with FLAG-SRSF10, the recovered material was fractionated, transferred on nitrocellulose that was decorated with anti-FLAG antibodies. “xx” and “x” indicate the large and small immunoglobulin subunits that react with the secondary antibody, respectively. Error bars indicate SD. Asterisks represent p values (two-tailed Student's t test) comparing the means between samples and their respective controls; *p < 0.05, **p < 0.01, and ***p < 0.001.