A single nucleotide in the SMN gene regulates splicing and is responsible for spinal muscular atrophy

Abstract

SMN1 and SMN2 (survival motor neuron) encode identical proteins. A critical question is why only the homozygous loss of SMN1, and not SMN2, results in spinal muscular atrophy (SMA). Analysis of transcripts from SMN1/SMN2 hybrid genes and a new SMN1 mutation showed a direct relationship between presence of disease and exon 7 skipping. We have reported previously that the exon-skipped product SMNDelta7 is partially defective for self-association and SMN self-oligomerization correlated with clinical severity. To evaluate systematically which of the five nucleotides that differ between SMN1 and SMN2 effect alternative splicing of exon 7, a series of SMN minigenes was engineered and transfected into cultured cells, and their transcripts were characterized. Of these nucleotide differences, the exon 7 C-to-T transition at codon 280, a translationally silent variance, was necessary and sufficient to dictate exon 7 alternative splicing. Thus, the failure of SMN2 to fully compensate for SMN1 and protect from SMA is due to a nucleotide exchange (C/T) that attenuates activity of an exonic enhancer. These findings demonstrate the molecular genetic basis for the nature and pathogenesis of SMA and illustrate a novel disease mechanism. Because individuals with SMA retain the SMN2 allele, therapy targeted at preventing exon 7 skipping could modify clinical outcome.

An in vivo reporter system for measuring increased inclusion of exon 7 in SMN2 mRNA: potential therapy of SMA

“…14,15 One of the nucleotides that differ creates a DdeI site in exon 8 of SMN2, thus RT-PCR analysis and digestion of the PCR products with DdeI distinguishes SMN1 and SMN2 expression. 16 By this analysis, the endogenous SMN1 and SMN2 mRNAs are equally expressed in essentially all cells and tissue types. The clue to the critical difference came from the observation that the transcripts undergo alternative splicing.…”

Emerging therapies and challenges in spinal muscular atrophy

“…Patients have variable copy numbers of the SMN2 gene, a related gene that differs from SMN1 by only five nucleotides, altering splicing and leading to transcription of a nonfunctional SMN protein lacking exon 7 in the majority of transcripts (Fig 1). 33 SMN2 copy number is the main determinant of phenotype, although not solely sufficient to predict severity 34. Sequence variations within the SMN2 gene and upregulation of modifier proteins, such as plastin 3, may also positively modify phenotype 35, 36, 37.…”

Baseline results of the NeuroNEXT spinal muscular atrophy infant biomarker study

“…SMA is caused by homozygous deletion or mutation in the SMN1 ( survival motor neuron 1 ) gene and retention of the nearly identical gene, SMN2 ( survival motor neuron 2 ), which results in reduced expression of full‐length SMN protein 3, 4. In humans, SMN2 is present in the same genomic region and differs from SMN1 by a single‐nucleotide substitution that results in the exclusion of exon 7 in approximately 90% of SMN transcripts 5, 6. The mRNA that results, SMNΔ7, produces a truncated protein that is nonfunctional and targeted for degradation 7, 8…”