Analysis of the 5-lipoxygenase promoter and characterization of a vitamin D receptor binding site.
1,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) and transforming growth factor beta (TGFbeta) potently induce 5-lipoxygenase (5-LO) in myeloid cells. We analyzed vitamin D receptor (VDR) binding to putative vitamin D response elements within the 5-LO promoter and analyzed its function by reporter gene analysis. Binding of VDR and retinoid X receptor to the promoter region was shown in DNase I footprinting, electrophoretic mobility shift and chromatin immunoprecipitation assays. However, the identified VDR binding region did not mediate induction of reporter gene activity by 1,25(OH)(2)D(3)/TGFbeta, neither in the 5-LO promoter context nor with the thymidine kinase (tk) promoter. Insertion of the rat atrial natriuretic factor VDRE in reporter plasmids containing the 5-LO promoter diminished induction by 1,25(OH)(2)D(3)/TGFbeta as compared with the tk promoter. Similarly, low inductions were obtained when cells were transiently or stably transfected with constructs containing various 5-LO promoter regions. Concerning basal promoter activity, we identified a positive regulatory region (-779 to -229), which includes the VDR binding region, in 5-LO-positive MonoMac6 cells. In summary, the VDR/RXR complex binds to putative VDREs in the 5-LO promoter, but other sequences outside the 5-LO promoter seem to be responsible or additionally required for the prominent induction of 5-LO mRNA expression by 1,25(OH)(2)D(3) and TGFbeta.
Sorg
Bernd L
BL
Institute of Pharmaceutical Chemistry/ZAFES, University of Frankfurt, Marie-Curie-Str. 9, D-60439 Frankfurt, Germany.
Klan
Niko
N
Seuter
Sabine
S
Dishart
David
D
Rådmark
Olof
O
Habenicht
Andreas
A
Carlberg
Carsten
C
Werz
Oliver
O
Steinhilber
Dieter
D
D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D002117:Calcitriol / Q000378:metabolism / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D017930:Genes, Reporter; D018922:HL-60 Cells; D006801:Humans; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D018167:Receptors, Calcitriol / Q000378:metabolism*; D047488:Retinoid X Receptors / Q000378:metabolism; D017421:Sequence Analysis; D017384:Sequence Deletion; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D020298:U937 Cells
D018167:Receptors, Calcitriol; D047488:Retinoid X Receptors; D016212:Transforming Growth Factor beta; D001094:Arachidonate 5-Lipoxygenase; D002117:Calcitriol