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

To investigate the impact of a novel heterozygous 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. : PBMC were isolated for mRNA analysis (cDNA synthesis, sequencing and one-step RT-PCR), Western Immunoblot (WI), and Immunofluorescence (IF). : cDNA analysis revealed the skipping of exon 15 and a premature stop codon at c.228. RT-PCR 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. : 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.