LAUSR

BACKGROUND AND PURPOSE Most spinal muscular atrophy (SMA) patients are homozygous for survival of motor neuron 1 gene (SMN1) deletion. However, some SMA patients carry an intragenic SMN1 mutation. Such patients provide a clue to understanding the function of the SMN protein and the role of each domain of the protein. We previously identified mutations in the Tudor domain and C-terminal region of the SMN protein in three Japanese SMA patients. To clarify the effect of these mutations on protein stability, we conducted expression assays of SMN with mutated domains. PATIENTS AND METHODS Patients A and B carried a mutation in SMN1 exon 3, which encodes a Tudor domain, c.275G>C (p.Trp92Ser). Patient C carried a mutation in SMN1 exon 6, which encodes a YG-box; c.819_820insT (p.Thr274Tyrfs). We constructed plasmid expression vectors containing wild-type and mutant SMN1 cDNAs. After transfection of HeLa cells with the expression plasmids, RNA and protein were isolated and analyzed by reverse-transcription PCR and western blot analysis. RESULTS The abundance of wild-type and mutant SMN1 transcripts in HeLa cells was almost the same. However, western blot analysis showed lower levels of mutant SMN proteins compared with wild-type SMN. In mutant SMN proteins, it is noteworthy that the level of the p.Thr274Tyrfs mutant was much reduced compared with that of the p.Trp92Ser mutant. CONCLUSIONS SMN mutations may affect the stability and levels of the protein.