Quantitative analysis of survival motor neuron copies: Identification of subtle SMN1 mutations in patients with spinal muscular atrophy, genotype-phenotype correlation, and implications for genetic counseling

Problems with diagnosis and genetic counseling occur for patients with auto somal recessive proximal spinal muscular atrophy (SMA) who do not show the most common mutation: homozygous absence of at least exon 7 of the telomeri c survival motor neuron gene (SMN1). Here we present molecular genetic data for 42 independent nondeleted SMA patients. A nonradioactive quantitative PCR test showed one SMN1 copy in 19 patients (45%). By sequencing cloned re verse-transcription (RT) PCR products or genomic fragments of SMN1, we iden tified nine different mutations in 18 of the 19 patients, six described for the first time: three missense mutations (Y272C, T274I, S262I), three fram eshift mutations in exons 2a, 2b, and 4 (124insT, 241-242ins4, 591delA), on e nonsense mutation in exon 1 (Q15X), one Alu-mediated deletion from intron 4 to intron 6, and one donor splice site mutation in intron 7 (c.922+6T--> G). The most frequent mutation, Y272C, was found in 6 (33%) of 18 patients. Each intragenic mutation found in at least two patients occurred on the sa me haplotype background, indicating founder mutations. Genotype-phenotype c orrelation allowed inference of the effect of each mutation on the function of the SMN1 protein and the role of the SMN2 copy number in modulating the SMA phenotype. In 14 of 23 SM4 patients with two SMN1 copies, at least one intact SMN1 copy was sequenced, which excludes a 5q-SMA and suggests the e xistence of further gene(s) responsible for similar to 4%-5% of phenotypes indistinguishable from SMA. We determined the validity of the test, and we discuss its practical implications and limitations.