The human T-cell leukemia virus type 1 (HTLV-1) transactivator (Tax) has been shown to interfere with regulated cellular proliferation. Many studies have focused on the ability of Tax to transform rodent fibroblasts; however, none has defined the molecular requirements for Tax transformation of human lymphoid cells. We show here that tax induces permanent growth of human primary T-lymphocytes by using a transformation/immortalization defective rhadinovirus vector. The cells phenotypically resemble HTLV-immortalized lymphocytes and contain episomally persisting recombinant rhadinoviral sequences, which stably express functional Tax protein. As Tax can activate major cellular signal transducing pathways including NF-B and cAMP-responsive element binding protein (CREB), we asked for the relevance of these routes in the immortalization of human lymphocytes. By using Tax mutants that either activate exclusively CREB/activating transcription factor or are defective in activating this signaling pathway, we delineated that Tax can induce immortalization of primary human T-lymphocytes through a mechanism independent of NF-B activation.

The human T-cell leukemia/lymphotropic virus type 1 (HTLV-1) induces a malignant lymphocytic disease. The HTLV-1 transactivator protein, Tax, is believed to be crucial for the development of the disease since it is transforming in vitro and induces tumors in transgenic animals. Although the transcriptional modulation of viral and cellular gene expression by Tax has been analyzed thoroughly, it has remained unclear how the Tax functions act on the cell cycle of primary T cells. To investigate the mechanism of Tax-mediated T-cell stimulation, we transduced primary human cord blood T cells with a conditional, tetracycline repressor-based tax expression system. Permanent Tax expression results in an abnormal proliferation of T cells which closely resemble HTLV-1-infected lymphocytes. Suppression of Tax synthesis stopped lymphocyte growth and caused cell cycle arrest in the G 1 phase. Upon reinduction of tax expression, the arrested cells entered the S phase. Thisshowed that Tax has mitogenic activity, which is required for stimulating the G 1 -to S-phase transition of immortalized lymphocytes. In mammalian cells, the G 1 -phase progression is controlled by the serial activation of several cyclin-dependent kinases (Cdks), starting with Cdk4 and Cdk6. In the presence of Tax, both Cdk4 and Cdk6 were activated. The suppression of Tax synthesis, however, resulted in a significant reduction of the Cdk4 and Cdk6 activities but did not influence the expression of Cdk4, Cdk6, or cognate D-type cyclin proteins. These data suggest that Tax induces Cdk4 and Cdk6 activity in primary human T lymphocytes; this Cdk activation is likely to account for the mitogenic Tax effect and for the abnormal T-cell proliferation of HTLV-1-infected lymphocytes.