The impact of paralog genes: detection of copy number variation in spinal muscle atrophy patients

Spinal muscular atrophy (SMA) is caused by dysfunction of the alpha motor neurons of the spinal cord. It is an autosomal recessive disease associated to the SMN1 gene, located in the subtelomeric region of 5q13. A paralog SMN2 gene is located at the centromeric region of the same chromosome, which apparently originated by an ancestral inverted duplication occurring only in humans. The exon sequence differs in two nucleotides in exon 7 and exon 8, which leads to an SMN2 transcript that lacks exon 7 and results in a truncated protein. Part (10%) of the SMN2 transcripts avoids the splicing of exon 7 but most of the copies are dysfunctional. In a disease scenario, the more SMN2 copies the higher possibility to restore at least partly the effects of SMN1 deficiency. Some therapeutic approaches are being developed to increase the expression of SMN2. To determine the number of SMN1 and SMN2 copies, the methodology must distinguish accurately between both genes. In this work, we present the results obtained using multiplex ligation-dependent probe amplification (MLPA) in 60 SMA suspected patients/carriers derived from different regions of Argentina. In 32 of these DNA samples we found alterations in SMN1. Among these, 16 presented a heterozygous deletion (carrier status) and 14 an homozygous deletion (patient status) in exon 7 and 8 of SMN1. In one case, exon 7 was found homozygously deleted but exon 8 presented a single copy, and in another case, exon 7 was found heterozygously deleted while exon 8 was normal. Almost half of the patients (7/15) presented a normal diploid number of SMN2 while the other half (8/15) presented an increased number. In this work we showed how a probe-based methodology such as MLPA was able to distinguish between the paralog genes and determine the amount of copies in DNA samples from suspected patients/carriers of SMA.