Nuclear translocation of SRPK2 and phosphorylation of SR proteins in response to PQ.

<p>A. Representative confocal micrographs of SH-SY5Y cells stained with an SRPK2-specific antibody. <i>Upper row</i>: cells treated with vehicle; <i>lower row</i>: cells treated with 0.75 mM PQ for 18 h. DAPI was used to identify the nuclei. B. The average nuclear to cytoplasmic ratio (N/C) ratio of SRPK2 fluorescent signal was determined for 50 cells as described in Materials and Methods. *** indicates p<0.001 treated vs. control group by unpaired t-test. C. RNA-mediated silencing strongly reduced SRPK1 and SRPK2 expression. Western blot analysis of the expression level of SRPK proteins in the same extract used for the Western blot shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0061980#pone-0061980-g002" target="_blank">figure 2D and 2E</a>. CPSF73K was used as loading control. D. Silencing of SRPK1 and SRPK2 abolishes PQ-mediated phosphorylation of SR proteins. Western blot analysis of nuclear extracts prepared from control SH-SY5Y cells treated with vehicle or with PQ, and from cells depleted of both SRPK1 and SRPK2. Phosphorylated SR proteins were detected with mab104. E. To control for equal loading of the samples SR proteins were also detected with mab16H3.