Protein synthesis is required for optimal induction of 25-hydroxyvitamin D(3)-24-hydroxylase, osteocalcin, and osteopontin mRNA by 1,25-dihydroxyvitamin D(3).

The regulation of the 25-hydroxyvitamin D(3)-24-hydroxylase gene by 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) has been extensively studied. It is well established that two vitamin D response elements in the promoter are responsible for the 1,25(OH)(2)D(3) induction of transcription. Surprisingly, this induction is blocked by the protein synthesis inhibitor, cycloheximide (CHX). In AOK-B50 cells, 1,25(OH)(2)D(3) caused a large induction of 24-hydroxylase mRNA by 7h; however, the addition of CHX simultaneously or 2h after 1,25(OH)(2)D(3) addition caused 76.4+/-13.0 and 37.1+/-18.8% reductions in the mRNA, respectively. Addition of CHX 4h after 1,25(OH)(2)D(3) had the opposite effect, and 21.7+/-17.2% more mRNA was observed after 7h. Similar patterns of mRNA expression were observed in other cell lines. CHX also decreased the induction by 1,25(OH)(2)D(3) of osteocalcin and osteopontin mRNA in ROS17/2.8 cells when added together with 1,25(OH)(2)D(3). The effect of CHX on the expression of a stably transfected luciferase construct under the control of 1400bp of 24-hydroxylase promoter indicates that a 1,25(OH)(2)D(3)-inducible transcription factor(s) that acts in the promoter region may at least in part be responsible for the effect of CHX on mRNA production of target genes.