A degron created by SMN2 exon 7 skipping is a principal contributor to spinal muscular atrophy severity

Howard Hughes Medical Institute and Department of Biochemistry and Biophysics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
Genes & development (Impact Factor: 10.8). 03/2010; 24(5):438-42. DOI: 10.1101/gad.1884910
Source: PubMed


Spinal muscular atrophy (SMA) is caused by homozygous survival of motor neurons 1 (SMN1) gene deletions, leaving a duplicate gene, SMN2, as the sole source of SMN protein. However, most of the mRNA produced from SMN2 pre-mRNA is exon 7-skipped ( approximately 80%), resulting in a highly unstable and almost undetectable protein (SMNDelta7). We show that this splicing defect creates a potent degradation signal (degron; SMNDelta7-DEG) at SMNDelta7's C-terminal 15 amino acids. The S270A mutation inactivates SMNDelta7-DEG, generating a stable SMNDelta7 that rescues viability of SMN-deleted cells. These findings explain a key aspect of the SMA disease mechanism, and suggest new treatment approaches based on interference with SMNDelta7-DEG activity.

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    • "However, although it encodes a virtually identical protein, the expression levels of SMN2 are not sufficient to restore full SMN activity (Arnold and Burghes, 2013; Nurputra et al., 2013). The coding regions of SMN1 and SMN2 differ only for a silent C to T transition at position 6 in exon 7, leading to the skipping of exon 7 in most SMN2 transcripts and to production of an unstable SMNΔ7 protein that is rapidly degraded (Burnett et al., 2009; Cho and Dreyfuss, 2010). The residual low levels of SMN2 transcripts that include exon 7 produce small amounts of fully functional SMN protein, supporting early development and viability in patients and mouse models of SMA (Arnold and Burghes, 2013; Nurputra et al., 2013). "
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    ABSTRACT: Spinal muscular atrophy (SMA) is a neurodegenerative disease caused by loss of motor neurons in patients with null mutations in the SMN1 gene. The almost identical SMN2 gene is unable to compensate for this deficiency because of the skipping of exon 7 during pre–messenger RNA (mRNA) processing. Although several splicing factors can modulate SMN2 splicing in vitro, the physiological regulators of this disease-causing event are unknown. We found that knockout of the splicing factor SAM68 partially rescued body weight and viability of SMAΔ7 mice. Ablation of SAM68 function promoted SMN2 splicing and expression in SMAΔ7 mice, correlating with amelioration of SMA-related defects in motor neurons and skeletal muscles. Mechanistically, SAM68 binds to SMN2 pre-mRNA, favoring recruitment of the splicing repressor hnRNP A1 and interfering with that of U2AF65 at the 3′ splice site of exon 7. These findings identify SAM68 as the first physiological regulator of SMN2 splicing in an SMA mouse model.
    Full-text · Article · Oct 2015 · The Journal of Cell Biology
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    • "Within SMN2, there is a C-to-T transition in an exonic splice enhancer of exon 7 that results in the vast majority (about 80–90%) of SMN2 mRNAs to lack exon 7 (SMNΔ7). SMNΔ7 is not fully functional and prone to degradation [11], [12]. SMN2 can, however, provide some fully functional SMN protein. "
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    ABSTRACT: Proximal spinal muscular atrophy (SMA) is an early onset, autosomal recessive motor neuron disease caused by loss of or mutation in SMN1 (survival motor neuron 1). Despite understanding the genetic basis underlying this disease, it is still not known why motor neurons (MNs) are selectively affected by the loss of the ubiquitously expressed SMN protein. Using a mouse embryonic stem cell (mESC) model for severe SMA, the RNA transcript profiles (transcriptomes) between control and severe SMA (SMN2+/+;mSmn-/-) mESC-derived MNs were compared in this study using massively parallel RNA sequencing (RNA-Seq). The MN differentiation efficiencies between control and severe SMA mESCs were similar. RNA-Seq analysis identified 3,094 upregulated and 6,964 downregulated transcripts in SMA mESC-derived MNs when compared against control cells. Pathway and network analysis of the differentially expressed RNA transcripts showed that pluripotency and cell proliferation transcripts were significantly increased in SMA MNs while transcripts related to neuronal development and activity were reduced. The differential expression of selected transcripts such as Crabp1, Crabp2 and Nkx2.2 was validated in a second mESC model for SMA as well as in the spinal cords of low copy SMN2 severe SMA mice. Furthermore, the levels of these selected transcripts were restored in high copy SMN2 rescue mouse spinal cords when compared against low copy SMN2 severe SMA mice. These findings suggest that SMN deficiency affects processes critical for normal development and maintenance of MNs.
    Full-text · Article · Sep 2014 · PLoS ONE
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    • "Most SMA cases (>96%) are due to mutations in the SMN1 gene, which encodes the survival motor neuron protein (Lefebvre et al. 1995). Humans have a second copy of this gene, termed SMN2, which is unaffected in SMA patients but cannot generally compensate for the SMN1 mutations because ∼80% of the SMN2 transcripts lack exon 7 and encode a protein isoform that is rapidly degraded (Cho and Dreyfuss 2010). Some SMA patients have additional copies of the SMN2 gene and display less severe symptoms, indicating that the severity of the disease directly correlates with the amount of functional SMN protein (Lefebvre et al. 1997). "
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    ABSTRACT: Spinal muscular atrophy is a neuromuscular disease resulting from mutations in the SMN1 gene, which encodes the survival motor neuron (SMN) protein. SMN is part of a large complex that is essential for the biogenesis of spliceosomal small nuclear RNPs. SMN also colocalizes with mRNAs in granules that are actively transported in neuronal processes, supporting the hypothesis that SMN is involved in axonal trafficking of mRNPs. Here, we have performed a genome-wide analysis of RNAs present in complexes containing the SMN protein and identified more than 200 mRNAs associated with SMN in differentiated NSC-34 motor neuron-like cells. Remarkably, ∼30% are described to localize in axons of different neuron types. In situ hybridization and immuno-fluorescence experiments performed on several candidates indicate that these mRNAs colocalize with the SMN protein in neurites and axons of differentiated NSC-34 cells. Moreover, they localize in cell processes in an SMN-dependent manner. Thus, low SMN levels might result in localization deficiencies of mRNAs required for axonogenesis.
    Preview · Article · Oct 2013 · RNA
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