1,25-Dihydroxyvitamin D3 enhances phorbol ester-stimulated differentiation and protein kinase C-dependent substrate phosphorylation activity in the U937 human monoblastoid cell.
In the U937 human monoblastoid cell line, 1,25-dihydroxyvitamin D3[1,25(OH)2D3] through a specific interaction with the 1,25(OH)2D3 receptor promotes differentiation toward a more mature phenotype. In addition to this direct effect, 1,25(OH)2D3 also potentiates differentiation in response to lymphokines and (Bu)2cAMP. We examined the effect of 1,25(OH)2D3 on phorbol ester-stimulated differentiation. Either preincubation with or simultaneous exposure to 1,25(OH)2D3 enhanced phorbol ester-stimulated differentiation. Over a 72-h period, the increase in phorbol ester responsiveness was dependent on the duration of 1,25(OH)2D3 exposure. Enhancement of phorbol ester-induced differentiation was observed with 1,25(OH)2D3 concentrations ranging from 0.1-10 nM. The 1,25(OH)2D3 vitamin D metabolite was more potent than the 24,25-dihydroxyvitamin D3 and 25-hydroxyvitamin D3 metabolites in potentiating phorbol ester-induced differentiation. Phorbol esters can exert at least a portion of their effects on cellular function by activating protein kinase C. Thus, one mechanism by which 1,25(OH)2D3 could amplify signal transduction leading to potentiation of phorbol ester-stimulated differentiation would be by enhancing phorbol ester-stimulated phosphorylation. To examine this possibility, we measured protein kinase C-dependent substrate phosphorylation in extracts derived from cells pretreated with 1,25(OH)2D3. In extracts derived from cells treated with 1,25(OH)2D3, the protein kinase C-dependent phosphorylation of endogenous U937 substrates stimulated by calcium, phosphatidyl serine, and diolein was increased compared to that observed in vehicle-treated cells. The conditions required for 1,25(OH)2D3 to increase protein kinase C-dependent phosphorylation of endogenous substrates (concentration, duration of exposure, and metabolite specificity) were similar to those required to enhance phorbol ester-stimulated differentiation. Possibly mediating this enhanced phosphorylation was an increase in protein kinase C activity observed in extracts derived from 1,25(OH)2D3-treated cells.
Ways
D K
DK
Department of Medicine, School of Medicine, East Carolina University, Greenville, North Carolina 27834-4354.
Dodd
R C
RC
Bennett
T E
TE
Gray
T K
TK
Earp
H S
HS
D002117:Calcitriol / Q000494:pharmacology*; D002448:Cell Adhesion / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D006801:Humans; D007700:Kinetics; D009363:Neoplasm Proteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D009363:Neoplasm Proteins; D011493:Protein Kinase C; D002117:Calcitriol; D013755:Tetradecanoylphorbol Acetate