Gene targeting by the vitamin D response element binding protein reveals a role for vitamin D in osteoblast mTOR signaling.
Transcriptional regulation by hormonal 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] involves occupancy of vitamin D response elements (VDREs) by the VDRE binding protein (VDRE-BP) or 1,25(OH)(2)D(3)-bound vitamin D receptor (VDR). This relationship is disrupted by elevated VDRE-BP, causing a form of hereditary vitamin D-resistant rickets (HVDRR). DNA array analysis showed that of 114 genes regulated by 1,25(OH)(2)D(3) in control cells, almost all (113) were rendered insensitive to the hormone in VDRE-BP-overexpressing HVDRR cells. Among these was the gene for DNA-damage-inducible transcript 4 (DDIT4), an inhibitor of mammalian target of rapamycin (mTOR) signaling. Chromatin immunoprecipitation PCR using 1,25(OH)(2)D(3)-treated osteoblasts confirmed that VDR and VDRE-BP compete for binding to the DDIT4 gene promoter. Expression of DDIT4 mRNA in these cells was induced (1.6-6 fold) by 1,25(OH)(2)D(3) (10-100 nM), and Western blot and flow cytometry analysis showed that this response involved suppression of phosphorylated S6K1(T389) (a downstream target of mTOR) similar to rapamycin treatment. siRNA knockdown of DDIT4 completely abrogated antiproliferative responses to 1,25(OH)(2)D(3), whereas overexpression of VDRE-BP exerted a dominant-negative effect on transcription of 1,25(OH)(2)D(3)-target genes. DDIT4, an inhibitor of mTOR signaling, is a direct target for 1,25(OH)(2)D(3) and VDRE-BP, and functions to suppress cell proliferation in response to vitamin D.
Orthopaedic Hospital Research Center, University of California-Los Angeles, 615 Charles E. Young Dr., South Los Angeles, CA 90095, USA.
Thomas S
TS
Lisse
Ting
T
Liu
Martin
M
Irmler
Johannes
J
Beckers
Hong
H
Chen
John S
JS
Adams
Martin
M
Hewison
D001859:Bone Neoplasms; D002117:Calcitriol / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology; D045744:Cell Line, Tumor; D053098:Familial Hypophosphatemic Rickets / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D006801:Humans; D010006:Osteoblasts / Q000166:cytology / Q000502:physiology; D012516:Osteosarcoma; D011401:Promoter Regions, Genetic / Q000502:physiology; D034741:RNA, Small Interfering; D018167:Receptors, Calcitriol / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology
C469991:DDIT4 protein, human; D034741:RNA, Small Interfering; D018167:Receptors, Calcitriol; D014157:Transcription Factors; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D002117:Calcitriol