REGULATION OF VIMENTIN EXPRESSION AND PROTEASE-MEDIATED VIMENTIN DEGRADATION DURING DIFFERENTIATION OF HUMAN MONOCYTIC LEUKEMIA-CELLS

Terminal differentiation of human monocytic leukemia THP-1 cells is in duced in vitro by 12-O-tetradenanoylphorbol-13-acetate (TPA). We inves tigated the effects of TPA on the expression of vimentin during the di fferentiation of THP-1 cells at both the mRNA and the protein level, O n northern blotting analysis, a 2.1 kb vimentin mRNA was up-regulated by TPA. On western blotting, small vimentin molecules with a molecular mass of approximately SO kDa were observed in the soluble fraction an d increased with TPA-induction of cellular differentiation. Since larg er, including intact, vimentin molecules were detectable at a high TPA dose, we assessed the possible existence of protease activity directe d against vimentin in THP-1 cells. With incubation of the cellular lys ates of THP-1 cells, the endogenous vimentin became increasingly small er over time, suggesting the presence of a vimentin-degrading protease . Phenylmethylsulfonyl fluoride inhibited this apparent protease activ ity against vimentin, suggesting the enzyme involved to be a serine pr otease, Interestingly, the protease activity was down-regulated by TPA treatment. TPA-treated TWP-1 cells were found to express a vimentin-f ilament network based on immunocytochemical analysis using an anti-vim entin monoclonal antibody, V9. Taken together, these observations sugg est that post-translational mechanisms work in cooperation with transc riptional regulation to maintain the vimentin-intermediate filament st ructure in differentiated THP-1 cells.