Nusinersen ameliorates motor function and prevents motoneuron Cajal body disassembly and abnormal poly(A) RNA distribution in a SMA mouse model

María T Berciano,Alba Puente-Bedia,José C Rodríguez-Rey,Olga Tapia,Jordi Calderó,Almudena Medina-Samamé,Miguel Lafarga

doi:10.1038/s41598-020-67569-3

Abstract

Spinal muscular atrophy (SMA) is a devastating autosomal recessive neuromuscular disease characterized by degeneration of spinal cord alpha motor neurons (αMNs). SMA is caused by the homozygous deletion or mutation of the survival motor neuron 1 (SMN1) gene, resulting in reduced expression of SMN protein, which leads to αMN degeneration and muscle atrophy. The majority of transcripts of a second gene (SMN2) generate an alternative spliced isoform that lacks exon 7 and produces a truncated nonfunctional form of SMN. A major function of SMN is the biogenesis of spliceosomal snRNPs, which are essential components of the pre-mRNA splicing machinery, the spliceosome. In recent years, new potential therapies have been developed to increase SMN levels, including treatment with antisense oligonucleotides (ASOs). The ASO-nusinersen (Spinraza) promotes the inclusion of exon 7 in SMN2 transcripts and notably enhances the production of full-length SMN in mouse models of SMA. In this work, we used the intracerebroventricular injection of nusinersen in the SMN∆7 mouse model of SMA to evaluate the effects of this ASO on the behavior of Cajal bodies (CBs), nuclear structures involved in spliceosomal snRNP biogenesis, and the cellular distribution of polyadenylated mRNAs in αMNs. The administration of nusinersen at postnatal day (P) 1 normalized SMN expression in the spinal cord but not in skeletal muscle, rescued the growth curve and improved motor behavior at P12 (late symptomatic stage). Importantly, this ASO recovered the number of canonical CBs in MNs, significantly reduced the abnormal accumulation of polyadenylated RNAs in nuclear granules, and normalized the expression of the pre-mRNAs encoding chondrolectin and choline acetyltransferase, two key factors for αMN homeostasis. We propose that the splicing modulatory function of nusinersen in SMA αMN is mediated by the rescue of CB biogenesis, resulting in enhanced polyadenylated pre-mRNA transcription and splicing and nuclear export of mature mRNAs for translation. Our results support that the selective restoration of SMN expression in the spinal cord has a beneficial impact not only on αMNs but also on skeletal myofibers. However, the rescue of SMN expression in muscle appears to be necessary for the complete recovery of motor function.

Highlights

Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disease that is considered the main genetic cause of infant mortality
It is well established that αMNs are the main physiopathological cellular targets of SMA, increasing evidence has revealed the primary importance of other peripheral tissues[41,57,58]
In our previous studies in the SMNΔ7 mouse model of SMA, we observed that SMN depletion in αMNs during the symptomatic postnatal stage severely impacts nuclear compartments involved in pre-rRNA and pre-mRNA processing[26]

Summary

Introduction

Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disease that is considered the main genetic cause of infant mortality. In addition to the SMN1 gene, humans ubiquitously express one or several copies of a closely related paralog gene called SMN2 This gene encodes a full-length SMN protein but presents one consistent difference, that is, the transition C to T (C6T) in exon 7. The physical association of CBs with the dense fibrillar component of the nucleolus, the site of synthesis and early processing of pre-rRNAs provides a molecular link for a nucleolus-CB interaction[18,22,23,24] In this context, recent studies from our laboratory have shown that reduced SMN levels in αMNs of a type I SMA patient and the SMN∆7 mouse model induce nuclear features of αMN degeneration, including depletion of canonical CBs, nucleolar stress and altered mRNA processing[25,26]. The dysfunction of RNA metabolism has enabled us to redefine the SMA as a RNA pathology, it could be considered a s pliceopathy[28]

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Results

Conclusion