Poster abstracts

Poster number 39 submitted by Fran Jodelka

Splicing of the spinal muscular atrophy-modifying gene, SMN2, is controlled by an auto-regulatory feedback loop.

Francine M. Jodelka (Department of Cell Biology and Anatomy, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL.), Allison D. Ebert (University of Wisconsin-Madison, Stem Cell and Regenerative Medicine Center, Waisman Center, Madison, WI.), Michelle L. Hastings (Department of Cell Biology and Anatomy, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL.)

Abstract:
Spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality. SMA is caused by mutations in the SMN1 gene, leading to motor neuron death and progressive muscle weakness and atrophy. SMN2 is nearly identical to SMN1 and both code for SMN protein. However, most of the mRNA produced from SMN2 lacks exon 7 and codes for a truncated, unstable protein product. SMN protein is an essential part of a complex that assembles Sm proteins onto spliceosomal snRNAs during the production of mature snRNPs that are required for splicing. We hypothesize that the loss of SMN1 in SMA leads to a reduction in snRNPs that consequently impacts SMN2 exon 7 splicing. We observe that high SMN protein levels correlate with greater SMN2 exon 7 splicing in mice and in induced pluripotent stem cells derived from a normal individual compared to cells from an SMA patient with lower SMN protein levels. These results suggest a link between SMN protein levels and exon 7 splicing, possibly via modulation of snRNP abundance. To test this idea, we inactivated individual snRNPs in vitro. We observed differential effects on exon 7 inclusion depending on which snRNP was targeted, suggesting that changes in snRNP levels affect exon 7 splicing. Altering U1 snRNP activity in cells using a binding-site decoy resulted in a decrease in endogenous SMN2 exon 7 inclusion, suggesting that U1 snRNP abundance affects exon 7 splicing. RNAi-mediated knockdown of SMN or Sm proteins resulted in a decrease in exon 7 splicing. Together, our results support the existence of a feedback loop in SMN expression by which low SMN protein levels exacerbate SMN exon 7 skipping, leading to a further reduction in SMN protein abundance. These results imply that an increase in SMN protein abundance may cause a disproportionately large increase in SMN expression, a finding that is important for understanding therapeutic potential in SMA treatment.

Keywords: RNA and disease, SMN, Spinal muscular atrophy