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. Author manuscript; available in PMC: 2015 Apr 1.
Published in final edited form as: Cancer Discov. 2014 Jul 11;4(10):1168–1181. doi: 10.1158/2159-8290.CD-13-0747

Figure 6.

Figure 6

The splicing regulator SRSF9 represses FGFR signaling through the cytoplasmic adaptor protein SH3BP2. A, Schematic of the RNA-Seq experimental approach and data analysis. B, qRT-PCR analysis monitoring expression of SRSF9, SH3BP2 exon 10 and SH3BP2 exons 7–8 (SH3BP2 total) in SA cells expressing a non-silencing (NS) shRNA or one of two unrelated SRSF9 shRNAs. The results were normalized to expression in NS control cells, which was set to 1. C, Schematic showing splicing of the SH3BP2 gene in NS and SRSF9 knockdown cells, in which exon 10 (yellow) is skipped. Red octagons indicate stop codons. The resulting proteins are also shown. In the truncated protein, the last two exons are shown in purple to indicate an alternate reading frame compared to the full-length protein. D, Immunoblot monitoring levels of SH3BP2 in SA cells expressing an NS or one of two SRSF9 shRNAs. E and F, Immunoblot analysis monitoring levels of tFGFR1 and SH3BP2 in SA cells expressing an SH3BP2 shRNA (E) or overexpressing SH3BP2 (F). G, Co-immunoprecipitation analysis. Left, the FGFR1 immunoprecipitate was immublotted for SH3BP2. Right, the SH3BP2 immunoprecipitate was immunoblotted for FGFR1. The levels of the proteins in the input are shown. H, Soft agar colony formation assay. NIH 3T3 cells expressing a NS shRNA or one of two unrelated SH3BP2 shRNAs were analyzed for their ability to form colonies in soft agar. I, Tumor formation in mice injected with NIH 3T3 cells expressing a NS shRNA or one of two unrelated SH3BP2 shRNAs. J, Model depicting the mechanism by which loss of SRSF9 leads to increased FGFR1 levels. K, Boxplots displaying log2 fold changes in expression of SH3BP2 exon 10 in normal lung samples, 27 hLSCC samples and 10 human lung adenocarcinoma (hLA) samples. Boxed areas span the first to the third quartile. Whiskers represent 15th and 85th percentiles. P<0.05 for hLSCC versus hLA. *P<0.05; **P<0.01.