Transfaction of avian vitamin D-dependent calbindin-D 28K 5' flanking promoter sequence in primary chick kidney cells

Abstract

Expression of the vitamin D induced calbindin-D 28K protein is transcriptionally controlled by the steroid hormone 1,25-dihydroxyvitamin D 3 (1, 25(OH) 2D 2) in a tissue-specific manner in the intestine and kidney. In order to examine the cis-acting elements of the calbindin-D 28Kpromoter and its modulation by 1,25-dihydroxyvitamin D 3, chimeric plasmids containing 2.1 kb of 5' flanking region linked to the reporter gene chloramphenicol acetyl transferase (CAT) were transfected by lipofection into primary cultures of chick kidney cells. Transfected chick kidney cells exhibited a high basal expression of the chloramphenicol acetyl transferase gene, reflecting the strong activity of the calbindin-D 28K promoter. Expression of the pCaBP2.1 reporter gene was increased 2-fold in the presence of the hormone 1,25(OH) 2D 3 in the primary kidney cells. Deletion of a 1.42 kb fragment ending —679 base pairs upstream from the transcription start site led to a 2-fold repression in the reporter gene activity by the hormone 1,25(OH) 2D 3 in primary chick kidney cultures. These preliminary results suggest that both positive and negative elements normally act to regulate the expression of the calbindin-D 28K gene in primary chick kidney cells.