Vitamin D receptor is not required for the rapid actions of 1,25-dihydroxyvitamin D3 to increase intracellular calcium and activate protein kinase C in mouse osteoblasts.
The rapid, non-genomic actions of 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] have been well described, however, the role of the nuclear vitamin D receptor (VDR) in this pathway remains unclear. To address this question, we used VDR(+/+) and VDR(-/-) osteoblasts isolated from wild-type and VDR null mice to study the increase in intracellular calcium ([Ca(2+)](i)) and activation of protein kinase C (PKC) induced by 1,25(OH)(2)D(3). Within 1 min of 1,25(OH)(2)D(3) (100 nM) treatment, an increase of 58 and 53 nM in [Ca(2+)](i) (n = 3) was detected in VDR(+/+) and VDR(-/-) cells, respectively. By 5 min, 1,25(OH)(2)D(3) caused a 2.1- and 1.9-fold increase (n = 6) in the phosphorylation of PKC substrate peptide acetylated-MBP(4-14) in VDR(+/+) and VDR(-/-) osteoblasts. The 1,25(OH)(2)D(3)-induced phosphorylation was abolished by GF109203X, a general PKC inhibitor, in both cell types, confirming that the secosteroid induced PKC activity. Moreover, 1,25(OH)(2)D(3) treatment resulted in the same degree of translocation of PKC-alpha and PKC-delta, but not of PKC-zeta, from cytosol to plasma membrane in both VDR(+/+) and VDR(-/-) cells. These experiments demonstrate that the 1,25(OH)(2)D(3)-induced rapid increases in [Ca(2+)](i) and PKC activity are neither mediated by, nor dependent upon, a functional nuclear VDR in mouse osteoblasts. Thus, VDR is not essential for these rapid actions of 1,25(OH)(2)D(3) in osteoblasts.
Wali
Ramesh K
RK
Evanston Northwestern Healthcare, Evanston, Illinois 60201, USA.
Kong
Juan
J
Sitrin
Michael D
MD
Bissonnette
Marc
M
Li
Yan Chun
YC
D000818:Animals; D002117:Calcitriol / Q000494:pharmacology / Q000502:physiology*; D002118:Calcium / Q000032:analysis / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007527:Isoenzymes / Q000096:biosynthesis; D008301:Maleimides / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D010006:Osteoblasts / Q000187:drug effects / Q000502:physiology*; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000032:analysis / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D012313:RNA / Q000032:analysis / Q000302:isolation & purification; D018167:Receptors, Calcitriol / Q000235:genetics / Q000502:physiology*
D004791:Enzyme Inhibitors; D007211:Indoles; D007527:Isoenzymes; D008301:Maleimides; D018167:Receptors, Calcitriol; D012313:RNA; D011493:Protein Kinase C; D002117:Calcitriol; C070515:bisindolylmaleimide I; D002118:Calcium