CYTOSOL IS THE PRIME COMPARTMENT OF HEPATITIS-B VIRUS-X PROTEIN WHEREIT COLOCALIZES WITH THE PROTEASOME

The hepatitis B virus X protein plays an important role in the regulat ion of viral genome expression and has also been implicated in the dev elopment of liver cancer associated with chronic viral infection. Seve ral effects have been attributed to X but their biological relevance r emains elusive. One of the confusing issues has been so far the uncert ainty concerning its cellular location. To gain insight into the mecha nism(s) how X exerts its effects, me have analysed its subcellular dis tribution and its dependency on the cell cycle. We used two complement ary approaches namely, immunolocalization using a cell line stably exp ressing X, and characterization of the dynamics of X location in livin g cells by means of the reporter gene GFP. Our data clearly define the cytosol as the prime location of X, irrespectively of the cell cycle and show in addition the close attachment of a fraction of X to the nu clear membrane. However, X does not associate with any cytoplasmic ves icles and organelles so far tested. In contrast, our study provides st rong evidence for the codistribution of X with the cytosolic fraction of proteasomes. In pulse-chase experiments, X decayed with a half-life of less than 30 min and proteasome-inhibitors did not modify its turn over, suggesting that X colocalization with the proteasome does not si mply point to its degradation pathway. The proteolytic processing of t he p105 precursor of the p50 subunit of the NF-kappa B transcription f actor, which has been shown to be proteasome-dependent, is markedly sl ow down in the presence of X. These findings suggest that X modulates the processing rate of p105 by acting presumably at the level of the p roteasome. Thus, targeting of proteasomes by X might be one of the pat hways employed by this viral protein to subvert pathways employed by c ellular functions.