Fig. 2.

Increased expression of HNRNPA1 in skeletal muscles promotes fetal splicing patterns for DM1 targets. (A) Systemic HNRNPA1 overexpression exacerbated, while MBNL2 partially reversed, DM1 fetal splicing patterns in HSA^LR muscles. RT-PCR analysis of Atp2a1 exon 22 and Ldb3 exon 11 splicing patterns were used as representative DM1 targets and positions of the fetal and adult exon splicing RT-PCR products are indicated (Right). Alternative cassette exon splicing was determined in wild-type (FVB) and HSA^LR mice without injection (control) or following AAV9-Mbnl2 (Mbnl2) or AAV9-HnrnpA1 HSA^LR injections (n = 4 each). (B) Immunoblotting for myc-tagged HNRNPA1 (αmyc), HNRNPA1 (αA1), CELF1 (αCELF1), and the loading control GAPDH (αGAPDH) following direct i.m. injections of AAV9-mycHnrnpa1 (n = 4 mice Hnrnp1.1-1.4), AAV9-mycMbnl1, or AAV9-mycMbnl2. The positions of myc-HNRNPA1 (myc-A1) and endogenous HNRNPA1 (A1) are indicated. (C) RT-PCR splicing analysis of six DM1 targeted gene transcripts demonstrated that HNRNPA1 overexpression in HSA^LR TA promoted fetal splicing events. The wild-type splicing pattern for each gene is shown in the FVB lanes. (D) Fetal exon inclusion was determined using percentage spliced in (Ψ; n = 4). P values were calculated using a one-way ANOVA with Tukey’s HSD post hoc test. Data are SEM and significant. ***P < 0.001.