Splicing of the Survival Motor Neuron genes and implications for treatment of SMA

Abstract

Proximal spinal muscular atrophy (SMA) is a neuromuscular disease caused by low levels of the survival motor neuron (SMN) protein. The reduced SMN levels are due to loss of the survival motor neuron-1 (SMN1) gene. Humans carry a nearly identical SMN2 gene that generates a truncated protein, due to a C to T nucleotide alteration in exon 7 that leads to inefficient RNA splicing of exon 7. This exclusion of SMN exon 7 is central to the onset of the SMA disease, however, this offers a unique therapeutic intervention in which corrective splicing of the SMN2 gene would restore SMN function. Exon 7 splicing is regulated by a number of exonic and intronic splicing regulatory sequences and trans-factors that bind them. A better understanding of the way SMN pre-mRNA is spliced has lead to the development of targeted therapies aimed at correcting SMN2 splicing. As therapeutics targeted toward correction of SMN2 splicing continue to be developed available SMA mouse models can be utilized in validating their potential in disease treatment.

Figure 1

Exon 7 ESE and ESS model of pre-mRNA splicing. Schematic of the SMN genes with each consisting of a weak 3’ and 5’ splice site. The SMN1 gene contains an ESE where SF2/ASF binding occurs which promotes exonic inclusion. In the SMN2 gene where position 6 contains a T nucleotide the SF2/ASF ESE is disrupted while a hnRNP A1 site is formed creating a new ESS. This mutated site disrupts exon recognition of exon 7 and promotes exon skipping.

Figure 2

SMN exon 7 is Regulated my multiple exonic and intronic regulators. Schematic of the SMN genes are shown with previously identified cis elements and any known proteins that bind them. Inclusion of exon 7 is regulated by the interaction of multiple factors.

Figure 3

Therapeutic strategies targeting splicing correction in SMN2. A. Standard ASOs targeting intron splicing silencers (ISS) and the exon 8 3'ss to promote exon 7 inclusion. B. Bifunctional ASOs linked to binding sites for positively acting splicing factors (SR proteins) to increase exon 7 inclusion. A bifunctional ASO linked to binding sites for inhibitory splicing factors (hnRNP proteins) to block the exon 8 3'ss and increase exon 7 inclusion by inhibiting splicing of exon 6 to 8. C. Trans splicing of an RNA molecule that can be used instead of SMN2 exon 7 to produce full length SMN transcripts. D. Drugs that promote the incorporation of exon 7 (green box) or inhibit the splicing of exon 6 to 8 (red box).