Wg. Chen et al., A NOVEL ARG(362)SER MUTATION IN THE STEROL 27-HYDROXYLASE GENE (CYP27) - ITS EFFECTS ON PRE-MESSENGER-RNA SPLICING AND ENZYME-ACTIVITY, Biochemistry (Easton), 37(43), 1998, pp. 15050-15056
A novel C to A mutation in the sterol 27-hydroxylase gene (CYP27) was
identified by sequencing amplified CYP27 gene products from a patient
with cerebrotendinous xanthomatosis (CTX). The mutation changed the ad
renodoxin cofactor binding residue (362)Arg to (362)Ser (CGT (362)Arg
to AGT (362)Ser), and was responsible for deficiency in the sterol 27-
hydroxylase activity, as confirmed by expression of mutant cDNA into C
OS-1 cells. Quantitative analysis showed that the expression of CYP27
gene mRNA in the patient represented 52.5% of the normal level. As the
mutation occurred at the penultimate nucleotide of exon 6 (-2 positio
n of exon 6-intron 6 splice site) of the gene, we hypothesized that th
e mutation may partially affect the normal splicing efficiency in exon
6 and cause alternative splicing elsewhere, which resulted in decreas
ed transcript in the patient. Transfection of constructed minigenes, w
ith or without the mutation, into COS-1 cells confirmed that the mutan
t minigene was responsible for a mRNA species alternatively spliced at
an activated cryptic 5' splice site 88 bp upstream from the 3' end of
exon 6. Our data suggest that the C to A mutation at the penultimate
nucleotide of exon 6 of the CYP27 gene not only causes the deficiency
in the sterol 27-hydroxylase activity, but also partially leads to alt
ernative pre-mRNA splicing of the gene. To our knowledge, this is the
first report regarding effects on pre-mRNA splicing of a mutation at t
he -2 position of a 5' splice site.