RT Journal Article
SR Electronic
T1 Full-length isoform sequencing for resolving the molecular basis of Charcot-Marie-Tooth 2A
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2023.02.07.526487
DO 10.1101/2023.02.07.526487
A1 Andrew B. Stergachis
A1 Elizabeth E. Blue
A1 Madelyn A Gillentine
A1 Lee-kai Wang
A1 Ulrike Schwarze
A1 Adriana Sedeño Cortés
A1 Jane Ranchalis
A1 Aimee Allworth
A1 Austin E. Bland
A1 Sirisak Chanprasert
A1 Jingheng Chen
A1 Daniel Doherty
A1 Andrew B. Folta
A1 Ian Glass
A1 Martha Horike-Pyne
A1 Alden Y. Huang
A1 Alyna T. Khan
A1 Kathleen A. Leppig
A1 Danny E. Miller
A1 Ghayda Mirzaa
A1 Azma Parhin
A1 Wendy Raskind
A1 Elisabeth A. Rosenthal
A1 Sam Sheppeard
A1 Samuel Strohbehn
A1 Virginia P. Sybert
A1 Thao T. Tran
A1 Mark Wener
A1 University of Washington Center for Mendelian Genomics (UW-CMG), Undiagnosed Diseases Network (UDN)
A1 Peter H. Byers
A1 Stanley F. Nelson
A1 Michael J. Bamshad
A1 Katrina M. Dipple
A1 Gail P. Jarvik
A1 Suzanne Hoppins
A1 Fuki M. Hisama
YR 2023
UL http://biorxiv.org/content/early/2023/02/07/2023.02.07.526487.abstract
AB Objectives Transcript sequencing of patient derived samples has been shown to improve the diagnostic yield for solving cases of likely Mendelian disorders, yet the added benefit of full-length long-read transcript sequencing is largely unexplored.Methods We applied short-read and full-length isoform cDNA 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 a branch point critical for intron 6 spicing. Full-length long-read isoform cDNA sequencing after treatment with a nonsense-mediated mRNA decay (NMD) inhibitor revealed that this variant creates five distinct altered splicing transcripts. All five 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 protein 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.Competing Interest StatementThe authors have declared no competing interest.