A novel splice site FUS mutation in a familial ALS case: effects on protein expression

Antonio Canosa, Annarosa Lomartire, Giovanni De Marco, Maurizio Grassano, Maura Brunetti, Umberto Manera, Rosario Vasta, Paolina Salamone, Giuseppe Fuda, Luca Sbaiz, Salvatore Gallone, Cristina Moglia (+2 others)
2021 figshare.com  
Objective: To investigate the impact of a novel heterozygous FUS mutation in the acceptor splice site of intron 14 (c.1542 − 1 g > t) on protein expression in Peripheral Blood Mononuclear Cells (PBMC) from a familial ALS patient. Methods: PBMC were isolated for mRNA analysis (cDNA synthesis, sequencing and one-step RT-PCR), Western Immunoblot (WI), and Immunofluorescence (IF). Results: cDNA analysis revealed the skipping of exon 15 and a premature stop codon at c.228. RT-PCR showed reduced FUS
more » ... showed reduced FUS mRNA by more than half compared to a healthy control (HC) and an ALS patient without genetic mutations (wtALS). In WI FUS band intensity in the proband was 30–50% compared to HC and wtALS. An antibody expected to detect only the wild-type protein did not reveal any reduction of FUS band intensity compared to the other antibodies. IF showed no difference among HC, wtALS, and the proband. Discussion: The reduction of FUS mRNA and protein in PBMC suggests the absence of the truncated protein, probably due to nonsense-mediated decay, leading to loss of function.
doi:10.6084/m9.figshare.14459021.v1 fatcat:ksi73lgwlvbonnp5oknkyqao6y