Transfection of avian vitamin D-dependent calbindin-D28K 5' flanking promoter sequence in primary chick kidney cells.
Expression of the vitamin D induced calbindin-D28K protein is transcriptionally controlled by the steroid hormone 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) in a tissue-specific manner in the intestine and kidney. In order to examine the cis-acting elements of the calbindin-D28K promoter and its modulation by 1,25-dihydroxyvitamin D3, 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-D28K promoter. Expression of the pCaBP2.1 reporter gene was increased 2-fold in the presence of the hormone 1,25(OH)2D3 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)2D3 in primary chick kidney cultures. These preliminary results suggest that both positive and negative elements normally act to regulate the expression of the calbindin-D28K gene in primary chick kidney cells.
Maiyar
A C
AC
Division of Biomedical Sciences, University of California, Riverside 92521.
Minghetti
P P
PP
Norman
A W
AW
D000818:Animals; D001483:Base Sequence; D064026:Calbindins; D002117:Calcitriol / Q000494:pharmacology; D002478:Cells, Cultured; D002645:Chickens; D005786:Gene Expression Regulation / Q000187:drug effects; D005796:Genes; D007668:Kidney / Q000378:metabolism* / Q000473:pathology; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D064030:S100 Calcium Binding Protein G / Q000096:biosynthesis / Q000235:genetics*; D014162:Transfection; D014808:Vitamin D Deficiency / Q000473:pathology
D064026:Calbindins; D011993:Recombinant Fusion Proteins; D064030:S100 Calcium Binding Protein G; D002117:Calcitriol