1,25-DIHYDROXYVITAMIN D-3 AND PHORBOL-MYRISTATE ACETATE SYNERGISTICALLY INCREASE CARBONIC ANHYDRASE-II EXPRESSION IN A HUMAN MYELOMONOCYTICCELL-LINE

Expression of carbonic anhydrase-II (CA-II), an enzyme important to os teoclast function, distinguishes osteoclasts from other cells of monoc ytic lineage. A cell's selection of terminal osteoclast phenotype is c ontrolled by many different factors, which are not well understood and which may also control the expression of CA-II. We studied the contro l of CA-II expression in the human HL-60 cell to better understand the signal transduction systems involved in progression to the osteoclast phenotype. Both 1,25-dihydroxyvitamin D-3 [1,25-(OH)(2)D-3; 10 nM] an d phorbol myristate acetate (10 ng/ml), doses that cause monocytic dif ferentiation of the HL-60 cell, induced small increases in CA-II mRNA and CA-II protein, as measured by Northern analysis and Western immuno blotting, respectively. The maximal response was seen at 3 days. Treat ment of HL-60 cells with both agents resulted in synergistic increases in CA-II mRNA (80-fold) and protein. The large increase in CA-II mRNA allowed assessment of the dose dependence of both agents, with ED(50) values of 1 nM for 1,25-(OH)(2)D-3 and 1 ng/ml for phorbol myristate acetate. In addition, we have shown that this synergistic response was completely inhibited by a potent inhibitor of protein kinase-C activi ty, staurosporine (0.1 mu M), which has not previously been demonstrat ed in other cell systems. Staurosporine did not inhibit 1,25-(OH)(2)D- 3 induction of nonspecific esterase. Thus, 1,25(OH)(2)D-3 synergistica lly interacts with protein kinase-C-activated systems to cause a myelo monocytic precursor to express CA-II, a marker of the osteoclast pheno type.