BACKGROUND: Spinal muscular atrophy (SMA) is an inherited neuromuscular disease affecting 1 in 8,000 newborns. The majority of patients carry bi-allelic variants in the survival of motor neuron 1 gene (SMN1). SMN1 is located in a duplicated region on chromosome 5q13 that contains Alu elements and is predisposed to genomic rearrangements. Due to the genomic complexity of the SMN region and genetic heterogeneity, approximately 50% of SMA patients remain without genetic diagnosis that is a prerequisite for genetic treatments. In this work we describe the diagnostic odyssey of one SMA patient in whom routine diagnostics identified only a maternal heterozygous SMN1Δ(7-8) deletion. METHODS: We characterized SMN transcripts, assessed SMN protein content in peripheral blood mononuclear cells (PBMC), estimated SMN genes dosage, and mapped genomic rearrangement in the SMN region. RESULTS: We identified an Alu-mediated deletion encompassing exons 2a-5 of SMN1 on the paternal allele and a complete deletion of SMN1 on the maternal allele as the cause of SMA in this patient. CONCLUSION: Alu-mediated rearrangements in SMN1 can escape routine diagnostic testing. Parallel analysis of SMN gene dosage, SMN transcripts, and total SMN protein levels in PBMC can identify genomic rearrangements and should be considered in genetically undefined SMA cases.

Department of Clinical Genetics Medirex A S Kosice Slovakia

Department of Medical Biology Faculty of Medicine P J Safarik University Kosice Slovakia

Research Unit for Rare Diseases Department of Pediatrics and Adolescent Medicine 1st Faculty of Medicine Charles University Prague Czech Republic

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Boczonadi, V. , Müller, J. S. , Pyle, A. , Munkley, J. , Dor, T. , Quartararo, J. , … Horvath, R. (2014). EXOSC8 mutations alter mRNA metabolism and cause hypomyelination with spinal muscular atrophy and cerebellar hypoplasia. Nature Communications, 5, 4287 10.1038/ncomms5287 PubMed DOI PMC

Butchbach, M. E. (2016). Copy number variations in the survival motor neuron genes: Implications for spinal muscular atrophy and other neurodegenerative diseases. Frontiers in Molecular Biosciences, 3, 7 10.3389/fmolb.2016.00007 PubMed DOI PMC

Cingolani, P. , Platts, A. , Wang, L. L. , Coon, M. , Nguyen, T. , Wang, L. , … Ruden, D. M. (2012). A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso‐2; iso‐3. Fly (Austin), 6(2), 80–92. 10.4161/fly.19695 PubMed DOI PMC

Crawford, T. O. , Paushkin, S. V. , Kobayashi, D. T. , Forrest, S. J. , Joyce, C. L. , Finkel, R. S. , … Chen, K. S. (2012). Evaluation of SMN protein, transcript, and copy number in the biomarkers for spinal muscular atrophy (BforSMA) clinical study. PLoS ONE, 7(4), e33572 10.1371/journal.pone.0033572 PubMed DOI PMC

Deininger, P. (2011). Alu elements: Know the SINEs. Genome Biology, 12(12), 236 10.1186/gb-2011-12-12-236 PubMed DOI PMC

Glascock, J. , Sampson, J. , Haidet‐Phillips, A. , Connolly, A. , Darras, B. , Day, J. , … Jarecki, J. (2018). Treatment algorithm for infants diagnosed with spinal muscular atrophy through newborn screening. Journal of Neuromuscular Diseases, 5(2), 145–158. 10.3233/JND-180304 PubMed DOI PMC

Groen, E. J. N. , Talbot, K. , & Gillingwater, T. H. (2018). Advances in therapy for spinal muscular atrophy: Promises and challenges. Nature Reviews Neurology, 14(4), 214–224. 10.1038/nrneurol.2018.4 PubMed DOI

Karakaya, M. , Paketci, C. , Altmueller, J. , Thiele, H. , Hoelker, I. , Yis, U. , & Wirth, B. (2019). Biallelic variant in AGTPBP1 causes infantile lower motor neuron degeneration and cerebellar atrophy. American Journal of Medical Genetics. Part A, 10.1002/ajmg.a.61198 PubMed DOI

Karakaya, M. , Storbeck, M. , Strathmann, E. A. , Delle Vedove, A. , Hölker, I. , Altmueller, J. , … Wirth, B. (2018). Targeted sequencing with expanded gene profile enables high diagnostic yield in non‐5q‐spinal muscular atrophies. Human Mutation, 39(9), 1284–1298. 10.1002/humu.23560 PubMed DOI

Kim, S. , Cho, C. S. , Han, K. , & Lee, J. (2016). Structural variation of Alu element and human disease. Genomics Information, 14(3), 70–77. 10.5808/GI.2016.14.3.70 PubMed DOI PMC

Lefebvre, S. , Bürglen, L. , Reboullet, S. , Clermont, O. , Burlet, P. , Viollet, L. , … Melki, J. (1995). Identification and characterization of a spinal muscular atrophy‐determining gene. Cell, 80(1), 155–165. 10.1016/0092-8674(95)90460-3 PubMed DOI

Livak, K. J. , Schmittgen, T. D. (2001). Analysis of relative gene expression data using real‐time quantitative PCR and the 2(‐Delta Delta C(T)) Method. Methods, 25(4), 402–408. 10.1006/meth.2001.1262 PubMed DOI

Majer F., Kousal B., Dusek P., Piherova L., Reboun M., Mihalova R., …, Sikora J. (2020). Alu‐mediated Xq24 deletion encompassing CUL4B, LAMP2, ATP1B4, TMEM255A, and ZBTB33 genes causes Danon disease in a female patient. American Journal of Medical Genetics Part A, 182(1), 219–223. PubMed

McKenna, A. , Hanna, M. , Banks, E. , Sivachenko, A. , Cibulskis, K. , Kernytsky, A. , … DePristo, M. A. (2010). The genome analysis toolkit: A MapReduce framework for analyzing next‐generation DNA sequencing data. Genome Research, 20(9), 1297–1303. 10.1101/gr.107524.110 PubMed DOI PMC

Mercuri, E. , Finkel, R. S. , Muntoni, F. , Wirth, B. , Montes, J. , Main, M. , … Szlagatys‐Sidorkiewicz, A. (2018). Diagnosis and management of spinal muscular atrophy: Part 1: Recommendations for diagnosis, rehabilitation, orthopedic and nutritional care. Neuromuscular Disorders, 28(2), 103–115. 10.1016/j.nmd.2017.11.005 PubMed DOI

Michelson, D. , Ciafaloni, E. , Ashwal, S. , Lewis, E. , Narayanaswami, P. , Oskoui, M. , & Armstrong, M. J. (2018). Evidence in focus: Nusinersen use in spinal muscular atrophy: Report of the Guideline Development, Dissemination, and Implementation Subcommittee of the American Academy of Neurology. Neurology, 91(20), 923–933. 10.1212/WNL.0000000000006502 PubMed DOI

Ottesen, E. W. , Seo, J. , Singh, N. N. , & Singh, R. N. (2017). A multilayered control of the human survival motor neuron gene expression by Alu elements. Frontiers in Microbiology, 8, 2252 10.3389/fmicb.2017.02252 PubMed DOI PMC

Paila, U. , Chapman, B. A. , Kirchner, R. , & Quinlan, A. R. (2013). GEMINI: Integrative exploration of genetic variation and genome annotations. PLoS Computational Biology, 9(7), e1003153 10.1371/journal.pcbi.1003153 PubMed DOI PMC

Renbaum, P. , Kellerman, E. , Jaron, R. , Geiger, D. , Segel, R. , Lee, M. , … Levy‐Lahad, E. (2009). Spinal muscular atrophy with pontocerebellar hypoplasia is caused by a mutation in the VRK1 gene. American Journal of Human Genetics, 85(2), 281–289. 10.1016/j.ajhg.2009.07.006 PubMed DOI PMC

Schorling, D. C. , Becker, J. , Pechmann, A. , Langer, T. , Wirth, B. , & Kirschner, J. (2019). Discrepancy in redetermination of SMN2 copy numbers in children with SMA. Neurology, 10.1212/WNL.0000000000007836 PubMed DOI

Shashi, V. , Magiera, M. M. , Klein, D. , Zaki, M. , Schoch, K. , Rudnik‐Schoneborn, S. , … Senderek, J. (2018). Loss of tubulin deglutamylase CCP1 causes infantile‐onset neurodegeneration. EMBO Journal, 37(23), 10.15252/embj.2018100540 PubMed DOI PMC

Song, X. , Beck, C. R. , Du, R. , Campbell, I. M. , Coban‐Akdemir, Z. , Gu, S. , … Lupski, J. R. (2018). Predicting human genes susceptible to genomic instability associated with Alu/Alu‐mediated rearrangements. Genome Research, 28(8), 1228–1242. 10.1101/gr.229401.117 PubMed DOI PMC

Stenson, P. D. , Mort, M. , Ball, E. V. , Shaw, K. , Phillips, A. , & Cooper, D. N. (2014). The human gene mutation database: Building a comprehensive mutation repository for clinical and molecular genetics, diagnostic testing and personalized genomic medicine. Human Genetics, 133(1), 1–9. 10.1007/s00439-013-1358-4 PubMed DOI PMC

Sumner, C. J. , Kolb, S. J. , Harmison, G. G. , Jeffries, N. O. , Schadt, K. , Finkel, R. S. , … Fischbeck, K. H. (2006). SMN mRNA and protein levels in peripheral blood: Biomarkers for SMA clinical trials. Neurology, 66(7), 1067–1073. 10.1212/01.wnl.0000201929.56928.13 PubMed DOI

Sun, Y. , Grimmler, M. , Schwarzer, V. , Schoenen, F. , Fischer, U. , & Wirth, B. (2005). Molecular and functional analysis of intragenic SMN1 mutations in patients with spinal muscular atrophy. Human Mutation, 25(1), 64–71. 10.1002/humu.20111 PubMed DOI

van der Steege, G. , Grootscholten, P. M. , Cobben, J. M. , Zappata, S. , Scheffer, H. , den Dunnen, J. T. , … Buys, C. H. (1996). Apparent gene conversions involving the SMN gene in the region of the spinal muscular atrophy locus on chromosome 5. American Journal of Human Genetics, 59(4), 834–838. Retrieved from http://www.ncbi.nlm.nih.gov/pubmed/8808598 PubMed PMC

Verhaart, I. E. C. , Robertson, A. , Wilson, I. J. , Aartsma‐Rus, A. , Cameron, S. , Jones, C. C. , … Lochmüller, H. (2017). Prevalence, incidence and carrier frequency of 5q‐linked spinal muscular atrophy ‐ a literature review. Orphanet Journal of Rare Diseases, 12(1), 124 10.1186/s13023-017-0671-8 PubMed DOI PMC

Wan, J. , Steffen, J. , Yourshaw, M. , Mamsa, H. , Andersen, E. , Rudnik‐Schöneborn, S. , … Jen, J. C. (2016). Loss of function of SLC25A46 causes lethal congenital pontocerebellar hypoplasia. Brain, 139(11), 2877–2890. 10.1093/brain/aww212 PubMed DOI PMC

Wan, J. , Yourshaw, M. , Mamsa, H. , Rudnik‐Schöneborn, S. , Menezes, M. P. , Hong, J. E. , … Jen, J. C. (2012). Mutations in the RNA exosome component gene EXOSC3 cause pontocerebellar hypoplasia and spinal motor neuron degeneration. Nature Genetics, 44(6), 704–708. 10.1038/ng.2254 PubMed DOI PMC

Wirth, B. , Herz, M. , Wetter, A. , Moskau, S. , Hahnen, E. , Rudnik‐Schöneborn, S. , … Zerres, K. (1999). 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. American Journal of Human Genetics, 64(5), 1340–1356. 10.1086/302369 PubMed DOI PMC

Easy and fast PCR-based protocol allows characterization of breakpoints resulting from Alu/Alu-mediated genomic rearrangements