The
enzyme 25-hydroxyvitamin D 1alpha-hydroxylase, or
CYP27B1, is the key
enzyme in the two-step activation process of
vitamin D to
1,25-dihydroxyvitamin D (1,25D). While a number of regulators of the renal
CYP27B1 enzyme activity have been recognized for some years, their underlying molecular mechanisms remain largely unknown, and the
DNA regions involved in the in vivo regulation of gene expression by these factors have not been delineated. We have generated a transgenic mouse line that expresses 1501 bp of 5' flanking region together with 44 bp of
5' untranslated region of the human
CYP27B1 gene fused to the
firefly luciferase reporter gene. Animals expressing the
luciferase gene demonstrated that both
luciferase protein and
mRNA for
CYP27B1 were localized to proximal convoluted tubule cells of the kidney. In 2-week-old animals, the expression of the transgene and the endogenous
CYP27B1 mRNA levels in the kidney were highest and fell with increasing age. Both reporter gene expression and
CYP27B1 mRNA levels were downregulated in response to increasing amounts of
dietary calcium in a dose-dependent manner.
Vitamin D deficiency resulted in an increase in both the reporter gene and
CYP27B1 expression. Interestingly, the increase in
CYP27B1 mRNA levels was substantially higher than the increase in reporter gene expression, suggesting either that there is a post-transcriptional mechanism that increases the amount of
CYP27B1 mRNA or that other regulatory elements are required to maximize the effect of
vitamin D deficiency. These findings demonstrate that the 1501 bp 5' flanking region of the
CYP27B1 gene directs expression to the proximal convoluted tubules of the kidney and is responsible for increasing transcriptional activity when
dietary calcium and
vitamin D levels are depleted. It also responds in the kidney to the physiological regulators of development and ageing.