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  • Phorbol esters enhance 1α,25-dihydroxyvitamin D3-regulated 25-hydroxyvitamin D-24-hydroxylase (CYP24A1) gene expression through ERK-mediated phosphorylation of specific protein 3 (Sp3) in Caco-2 cells.

Phorbol esters enhance 1α,25-dihydroxyvitamin D3-regulated 25-hydroxyvitamin D-24-hydroxylase (CYP24A1) gene expression through ERK-mediated phosphorylation of specific protein 3 (Sp3) in Caco-2 cells.

Molecular and cellular endocrinology (2012-08-09)
Yan Jiang, James C Fleet
PMID22871965
ABSTRACT

Phorbol 12-myristate 13-acetate (PMA) increased 1,25(OH)(2)D(3)-induced human 25 hydroxyvitamin d-24 hydroxylase (hCYP24A1) gene expression and vitamin D receptor (VDR) binding to the hCYP24A1 promoter. It did not alter transient receptor potential cation channel, subfamily V, member 6 (TRPV6) expression, VDR binding to the TRPV6 promoter, or VDR binding to a crude chromatin preparation. PMA activated Extracellular signal-Regulated Kinases (ERK) 1/2 and p38 mitogen activated protein kinases (MAPK) and inhibiting these kinases reduced 1,25(OH)(2)D(3)-induced and PMA-enhanced hCYP24A1 promoter activity. Mithramycin A inhibits Specific Protein (Sp) family member binding to DNA and reduced 1,25(OH)(2)D(3)-induced and PMA-enhanced hCYP24A1 promoter activity. Sp1 or Sp3 siRNA knockdown reduced 1,25(OH)(2)D(3)-regulated hCYP24A1 promoter activity but only Sp3 siRNA reduced PMA-enhanced hCYP24A1 promoter activity. PMA increased MAPK-dependent Sp3 phosphorylation, Sp3-VDR interactions, and Sp3 binding to the hCYP24A1 promoter. These data suggest that MAPK signaling contributes to 1,25(OH)(2)D(3)-induced and PMA-enhanced CYP24A1 gene transcription by modulating Sp3 function.

MATERIALS
Product Number
Brand
Product Description
524400
Sigma-Aldrich
Phorbol-12-myristate-13-acetate, Phorbol-12-myristate-13-acetate, CAS 16561-29-8, is the most common phorbol ester. Activates PKC at nanomolar concentrations.
662005
Sigma-Aldrich
U0126, U0126, CAS 109511-58-2, is a potent and specific inhibitor of MEK1 (IC50 = 72 nM) and MEK2 (IC50 = 58 nM). The inhibition is noncompetitive with respect to both ATP and ERK.
M6891
Sigma-Aldrich
Mithramycin A from Streptomyces plicatus, ≥90% (HPLC)
365250
Sigma-Aldrich
Gö 6976, Gö 6976, CAS 136194-77-9, is a cell-permeable, reversible, and ATP-competitive inhibitor of PKC (IC50 = 7.9 nM for rat brain). Exhibits selectively for PKCα (IC50 = 2.3 nM) and βI (IC50 = 6.2 nM).