ExSpeU1 rescue exonic mutations in FIX exon 5 through an <i>SRSF2</i>-mediated mechanism.
2016-05-26T20:26:53Z (GMT) by
<p>(A) Schematic representation of the secondary structure of ExSpeU1 RNA highlighting the 5' tail region (FIX). FIX exon 5 (uppercase) and intron 6 (lowercase) sequences are indicated along with the ExSpeU1 binding regions. (B) Cotransfection experiments. The R116R minigene was transfected in HeLa cells alone (NT) or with the indicated ExSpeU1s and the resulting splicing pattern evaluated by RT-PCR. (C) Splicing rescue activity mediated by ExSpeU1 FIX9 on exon skipping mutations. The indicated minigenes were co-transfected with (+) or without (-) ExSpeU1 FIX9 and the pattern of splicing evaluated. All data are the average of 3 replicates with error bars indicating SD. (D) ExSpeU1 rescue depends on SRSF2. siRNA treated (+) and control HeLa cells (-) were transfected with R116R minigene in the presence (lanes 3 and 4) or absence (lanes 1 and 2) of ExSpeU1. Data represent the average of 3 replicates with error bars indicating SD. (E) <i>SRSF2</i> and ExSpeU1 act regardless of endogenous U1snRNP. Wt and R116R minigene were transfected alone (lane 1 and 7, respectively) or in the presence of the indicated plasmids. D1 is antisense to the endogenous snRNA, whereas D3 is the control that contains a mutation that prevents binding to U1. In all panels data are the average of 3 replicates with error bars indicating SD.</p>