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5-Lipoxygenase (5-LO; EC 1.13.11.34) activity in the human monocytic cell line Mono Mac 6 was upregulated by combined treatment with transforming growth factor beta 1 (TGF-beta) and 1,25-dihydroxyvitamin D3 (VD3). In undifferentiated cells, 5-LO enzyme activity was undetectable. After the addition of TGF-beta plus VD3, the activity of intact cells was 800 ng per 10(6) cells--500 times more than the assay detection limit. Also 5-LO protein and mRNA expression were induced &gt; 128-fold and 64-fold, respectively, as compared to undifferentiated cells. Both TGF-beta and VD3 were required for these prominent responses. Either agent alone gave small amounts of 5-LO protein and mRNA but very low 5-LO activities. After the addition of TGF-beta and VD3, the induction of 5-LO protein was obvious after 1 day, but the increase in activity was delayed and did not appear until the second day. Pretreatment of cells with TGF-beta or VD3 alone for 2 days led to 5-LO protein expression but very low enzyme activity. Addition of the lacking second inducer was required for full induction of 5-LO protein expression and for upregulation of enzyme activity. Partial purification of 5-LO from Mono Mac 6 cells and recombination with soluble cellular proteins from different sources indicated the presence of cytosolic factors that affect the activity of 5-LO.

proceedings of the national academy of sciences of the united states of america

Sequential induction of 5-lipoxygenase gene expression and activity in Mono Mac 6 cells by transforming growth factor beta and 1,25-dihydroxyvitamin D3.