A Single Nucleotide Difference That Alters Splicing Patterns Distinguishes the SMA Gene SMN1 From the Copy Gene SMN2

Spinal muscular atrophy (SMA) is a recessive disorder characterized by loss of motor neurons in the spinalcord. It is caused by mutations in the telomeric survival motor neuron 1 (SMN1) gene. Alterations within analmost identical copy gene, the centromeric survival motor neuron 2 (SMN2) gene produce no known pheno-typic effect. The exons of the two genes differ by just two nucleotides, neither of which alters the encodedamino acids. At the genomic level, only five nucleotides that differentiate the two genes from one anotherhave been reported. The entire genomic sequence of the two genes has not been determined. Thus, differ-ences which might explain why SMN1is the SMA gene are not readily apparent. In this study, we have com-pletely sequenced and compared genomic clones containing the SMNgenes. The two genes show strikingsimilarity, with the homology being unprecedented between two different yet functional genes. The only crit-ical difference in an ~32 kb region between the two SMNgenes is the C→→→→T base change 6 bp inside exon 7.This alteration but not other variations in the SMNgenes affects the splicing pattern of the genes. The majorityof the transcript from the SMN1locus is full length, whereas the majority of the transcript produced by theSMN2locus lacks exon 7. We suggest that the exon 7 nucleotide change affects the activity of an exon spliceenhancer. In SMA patients, the loss of SMN1but the presence of SMN2results in low levels of full-lengthSMNtranscript and therefore low SMN protein levels which causes SMA.INTRODUCTIONProximal spinal muscular atrophy (SMA) is an autosomalrecessive neuromuscular disorder characterized by destructionof motor neurons in the anterior horn of the spinal cord. SMAhas an estimated incidence of 1 in 10 000 live births, with a car-rier frequency of ~1 in 50 people (1). Childhood onset SMA isclassified into three groups on the basis of age at onset andclinical course (2); type I SMA (Werdnig–Hoffman disease) isthe most severe form, with onset before the age of 6 monthsand death usually occurring within the first 2 years. Type IISMA is intermediate in severity. Onset occurs at ~18 monthsand patients never gain the ability to walk. Type III SMA(Kugelberg–Welander disease) is the mildest form of the dis-ease with onset after 18 months. Type III patients are able tostand and walk.All three forms of proximal SMA are due to mutations in thetelomeric but not centromeric survival motor neuron (SMN)genes (3–11). The full-length cDNAs of the two genes areidentical except for single nucleotide differences in exons 7and 8, yet their transcriptional products are not the same.SMN1 produces a majority of the full-length cDNA;SMN2produces mostly transcript lacking exon 7 (3). We have shownpreviously that promoter differences do not account for the dif-ferent levels of full-length transcript from the two genes (12).Instead, the exon 7 difference between the two genes affectssplicing, causing increased levels of full-length transcript fromSMN1 as compared with SMN2 (13).The SMN protein is a 38 kDa polypeptide which is ubiqui-tously expressed (14,15). It is found at especially high levels inthe spinal motor neurons. The exact function of the proteinremains unknown. However, recent studies have implicated itsinvolvement in mRNA biogenesis. Specifically, SMN has been