IDENTIFICATION OF A NEGATIVE ELEMENT IN THE HUMAN VIMENTIN PROMOTER - MODULATION BY THE HUMAN T-CELL LEUKEMIA-VIRUS TYPE-I TAX PROTEIN

The vimentin gene is a member of the intermediate filament multigene family and encodes a protein expressed, in vivo, in all mesenchymal derivatives and, in vitro, in cell types of various origin. We have previously demonstrated that the expression of this growth-regulated gene could be trans activated by the 40-kDa Tax protein of HTLV-I (human T-cell leukemia virus type I) and that responsiveness to this viral protein was mediated by the presence of an NF-kappaB binding site located between -241 and -210 bp upstream of the mRNA cap site (A. Lilienbaum, M. Duc Dodon, C. Alexandre, L. Gazzolo, and D. Paulin, J. Virol. 64:256-263, 1990). These previous assays, performed with deletion mutants of the vimentin promoter linked to the chloramphenicol acetyltransferase gene, also revealed the presence of an upstream negative region between -529 and -241 bp. Interestingly, the inhibitory activity exerted by this negative region was overcome after cotransfection of a Tax-expressing plasmid. In this study, we further characterize the vimentin negative element and define the effect of the Tax protein on the inhibitory activity of this element. We first demonstrate that a 187-bp domain (-424 to -237 bp) behaves as a negative region when placed upstream either of the NF-kappaB binding, site of vimentin or of a heterologous enhancer such as that present in the desmin gene promoter. The negative effect can be further assigned to a 32-bp element which is indeed shown to repress the basal or induced activity of the NF-kappaB binding site. Indeed, the negative effect is still observed in HeLa cells treated with phorbol myristate acetate, an inducer of nuclear NF-kappaB activity. Furthermore, deletion experiments allowed identification within the NE site of 19 bp which bind in vitro a protein complex and which are essential to exert the negative effect. We further demonstrate that expression of the Tax protein is required for overcoming the down-regulation of the NF-kappaB binding site by the negative element. Indeed, the negative effect can be relieved in HeLa cells cotransfected with a Tax-expressing plasmid or in a clone of HeLa cells stably producing this viral protein. Collectively, these observations give a new insight into the complex regulation of the human vimentin gene expression and suggest that the Tax protein of human T-cell leukemia virus type I up-regulates this cellular promoter through a dual mechanism consisting of the activation of the NF-kappaB binding site and the relief of the negative effect exerted by the negative element.