THE HEPATITIS-B VIRUS-X PROTEIN INCREASES THE CELLULAR-LEVEL OF TATA-BINDING PROTEIN, WHICH MEDIATES TRANSACTIVATION OF RNA-POLYMERASE-III GENES

The hepatitis B virus X gene product transactivates a variety of cellu lar and viral genes. The mechanism for X induction of RNA polymerase ( pol) III genes was investigated, By using Drosophila S-2 cells stably transformed with the X gene, the transient expression of a tRNA gene i s enhanced. Comparing the transcriptional activities of extracts deriv ed from these cells, all three types of RNA pol III promoters are stim ulated by X. Interestingly, both S-2 and rat 1A cells stably transform ed with the X gene produce increased cellular levels of the TATA-bindi ng protein (TBP). By using various kinase inhibitors, it was found tha t the X-mediated increases in both transcription and TBP are dependent upon protein kinase C activation, Since TBP is a subunit of TFIIIB, t he activity of this component fractionated from extracts derived from control and X-transformed cells was analyzed. These studies reveal tha t TFIIIB activity is substantially more limiting in control cells and that TFIIIB isolated from X-transformed cells has increased activity i n reconstitution assays compared with TFIIIB isolated from control cel ls. Conversely, comparison of TFIIIC from control and X-transformed ce ll extracts revealed that there is relatively little change in its abi lity either to reconstitute transcription or to bind to DNA and that t here is no change in the catalytic activity of RNA pol III, Studies we re performed to determine whether directly increasing cellular TBP alo ne could enhance RNA pol III gene transcription. Transient expression of a TBP cDNA in rat 1A cells was capable of stimulating transcription activity from the resultant extracts in vitro. Together, these result s demonstrate that one mechanism by which X mediates transactivation o f RNA pol III genes is by increasing limiting TBP via the activation o f cellular signaling pathways. The discovery that X increases cellular TBP, the universal transcription factor, provides a novel mechanism f or the function of a viral transactivator protein and may explain the ability of X to produce such large and diverse effects on cellular gen e expression.