INTERACTION OF THE HUMAN T-CELL LYMPHOTROPIC VIRUS TYPE-1 TAX TRANSACTIVATOR WITH TRANSCRIPTION FACTOR IIA

The Tax protein of human T-cell lymphotropic virus type 1 (HTLV-1) is a 40-kDa transcriptional activator which is critical for HTLV-1 gene r egulation and virus-induced cellular transformation. Tax is localized to the DNA through its interaction with the site-specific activators c yclic AMP-responsive element-binding protein, NF-kappa B, and serum re sponse factor. It has been suggested that the recruitment of Tax to th e DNA positions Tax for interaction with the basal transcriptional mac hinery. On the basis of several independent assays, we now report a ph ysical and functional interaction between Tax and the transcription fa ctor, TFIIA. First, Tax was found to interact with the 35-kDa (alpha) subunit of TFIIA in the yeast two-hybrid interaction system. Important ly, two previously characterized mutants with point mutations in Tax, M32 (Y196A, K197S) and M41 (N287A, P288S), which were shown to be defe ctive in Tax-activated transcription were unable to interact with TFII A in this assay. Second, a glutathione-S-transferase (GST) affinity-bi nding assay showed that the interaction of holo-TFIIA with GST-Tax was 20-fold higher than that observed with either the GST-Tax M32 activat ion mutant or the GST control. Third, a coimmunoprecipitation assay sh owed that in HTLV-1-infected human T lymphocytes, Tax and TFIIA were a ssociated. Finally, TFIIA facilitates Tax transactivation in vitro and in vivo. In vitro transcription studies showed reduced levels of Tax- activated transcription in cell extracts depleted of TFIIA. In additio n, transfection of human T lymphocytes with TFIIA expression vectors e nhanced Tax-activated transcription of an HTLV-1 long terminal repeat- chloramphenicol acetyltransferase reporter construct. Our study sugges ts that the interaction of Tax with the transcription factor TFIIA may play a role in Tax-mediated transcriptional activation.