1,25-Dihydroxyvitamin D3 translocates protein kinase C beta to nucleus and enhances plasma membrane association of protein kinase C alpha in renal epithelial cells.
1,25-Dihydroxycholecalciferol (1,25-(OH)2-D3) increases membrane-associated protein kinase C (PKC) activity and immunoreactivity in renal epithelial (Madin Darby bovine kidney, MDBK) cells (Simboli-Campbell, M., Franks, D. J., and Welsh, J. E. (1992) Cell Signalling 4, 99-109). We have now characterized the effects of 1,25-(OH)2-D3 on the subcellular localization of three individual isozymes by immunofluorescence and immunoblotting. Although the total amount of PKC alpha, PKC beta, and PKC zeta are unaffected by 1,25-(OH)2-D3, this steroid hormone induces subcellular redistribution of both PKC alpha and PKC beta. Treatment with 1,25-(OH)2-D3 (100 nM, 24 h) enhances plasma membrane association of PKC alpha and induces translocation of PKC beta to the nuclear membrane. The effects of 1,25-(OH)2-D3 appear to be limited to the calcium-dependent PKC isozymes, since 1,25-(OH)2-D3 has no effect on the calcium independent isozyme, PKC zeta. In contrast to rapid transient PKC translocation seen in response to agents which interact with membrane receptors to induce phospholipid hydrolysis, modulation of PKC alpha and PKC beta is observed after 24 h treatment with 1,25-(OH)2-D3. In MDBK cells, the phorbol ester 12-0-tetradecanoylphorbol-13-acetate (TPA) (100 nM, 24 h) down-regulates PKC alpha and, to a lesser extent, PKC zeta, without altering their subcellular distribution. TPA also induces translocation of PKC beta to the nuclear membrane. MDBK cells treated with 1,25-(OH)2-D3, but not TPA, exhibit enhanced phosphorylation of endogenous nuclear proteins. In addition to the distinct effects of 1,25-(OH)2-D3 and TPA on PKC isozyme patterns, 1,25-(OH)2-D3 up-regulates both the vitamin D receptor and calbindin D-28K, whereas TPA down-regulates the expression of both proteins. These data support the involvement of PKC in the mechanism of action of 1,25-(OH)2-D3 and specifically implicate PKC beta in 1,25-(OH)2-D3-mediated nuclear events.
Simboli-Campbell
M
M
Department of Biochemistry, University of Ottawa, Ontario, Canada.
Gagnon
A
A
Franks
D J
DJ
Welsh
J
J
D000818:Animals; D064026:Calbindins; D002117:Calcitriol / Q000494:pharmacology*; D002417:Cattle; D002458:Cell Fractionation; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology*; D002467:Cell Nucleus / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004848:Epithelium / Q000201:enzymology; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D007527:Isoenzymes / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D007668:Kidney / Q000201:enzymology*; D011493:Protein Kinase C / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D018167:Receptors, Calcitriol / Q000187:drug effects / Q000378:metabolism; D064030:S100 Calcium Binding Protein G / Q000032:analysis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D064026:Calbindins; D007527:Isoenzymes; D018167:Receptors, Calcitriol; D064030:S100 Calcium Binding Protein G; D011493:Protein Kinase C; D002117:Calcitriol; D013755:Tetradecanoylphorbol Acetate