Objectives: Transcript sequencing of patient-derived samples has been shown to improve the diagnostic yield for solving cases of suspected Mendelian conditions, yet the added benefit of full-length long-read transcript sequencing is largely unexplored.
Methods: We applied short-read and full-length transcript sequencing and mitochondrial functional studies to a patient-derived fibroblast cell line from an individual with neuropathy that previously lacked a molecular diagnosis.
Results: We identified an intronic homozygous MFN2 c.600-31T>G variant that disrupts the branch point critical for intron 6 splicing. Full-length long-read isoform complementary DNA (cDNA) sequencing after treatment with a nonsense-mediated mRNA decay (NMD) inhibitor revealed that this variant creates 5 distinct altered splicing transcripts. All 5 altered splicing transcripts have disrupted open reading frames and are subject to NMD. Furthermore, a patient-derived fibroblast line demonstrated abnormal lipid droplet formation, consistent with MFN2 dysfunction. Although correctly spliced full-length MFN2 transcripts are still produced, this branch point variant results in deficient MFN2 levels and autosomal recessive Charcot-Marie-Tooth disease, axonal, type 2A (CMT2A).
Discussion: This case highlights the utility of full-length isoform sequencing for characterizing the molecular mechanism of undiagnosed rare diseases and expands our understanding of the genetic basis for CMT2A.
Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.