Isolation and characterization of the rat alpha 1(I) collagen promoter. Regulation by 1,25-dihydroxyvitamin D.
Our previous work demonstrated that the inhibition of type I collagen synthesis by 1,25-dihydroxyvitamin D (1,25-(OH)2D3) in fetal rat calvaria and cultured rat osteosarcoma cells is accompanied by equivalent reduction in steady state levels of alpha 1(I) and alpha 2(I) collagen mRNA. To pursue the mechanism for this effect, we isolated and sequenced a 3.6-kilobase DNA fragment that contained the promoter for the rat alpha 1(I) collagen gene. This promoter fragment was fused to the chloramphenicol acetyltransferase gene and was introduced into ROS 17/2.8 cells by calcium phosphate co-precipitation. Expression of this construct was diminished by 1,25-(OH)2D3 to the same degree as the endogenous collagen gene in both transient expression assays and in permanently selected bone cells. However, a fibroblast cell line did not show a similar reduction in the activity of the transgene or the endogenous collagen gene. These experiments indicate that the alpha 1(I) promoter contains cis-active elements which are regulated by the 1,25-(OH)2D3 receptor in ROS 17/2.8 cells.
Lichtler
A
A
Department of Pediatrics, University of Connecticut Health Center, Farmington 06032.
Stover
M L
ML
Angilly
J
J
Kream
B
B
Rowe
D W
DW
D000818:Animals; D001483:Base Sequence; D002117:Calcitriol / Q000494:pharmacology*; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003001:Cloning, Molecular; D003094:Collagen / Q000235:genetics*; D015342:DNA Probes; D004262:DNA Restriction Enzymes; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D051379:Mice; D008969:Molecular Sequence Data; D012516:Osteosarcoma / Q000378:metabolism; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D014162:Transfection; D014407:Tumor Cells, Cultured
D015342:DNA Probes; D012333:RNA, Messenger; D003094:Collagen; D015500:Chloramphenicol O-Acetyltransferase; D004262:DNA Restriction Enzymes; D002117:Calcitriol