The hepatitis B virus X protein is a promiscuous transcriptional transactivator. Transactivation by the X protein is most likely mediated through binding to different cellular factors. Using the yeast two-hybrid method, we have isolated a clone that encodes a novel X-associated cellular protein: XAP2. X and XAP2 interactions also occur in vitro. Antiserum raised against XAP2 recognizes a cytoplasmic protein with an apparent molecular mass of 36 kDa. The interaction between X and XAP2 requires a small region on X containing amino acids 13-26. From Northern blot analyses, XAP2 is ubiquitously expressed in both liver-derived and non-liver-derived cell lines as well as in normal non-liver tissues. In contrast, XAP2 is expressed in very low level in the normal human liver. In transfection assays, overexpression of XAP2 abolishes transactivation by the X protein. Based on these results, we suggest that XAP2 is an important cellular negative regulator of the X protein, and that X-XAP2 interaction may play a role in HBV pathology.

The hepatitis B virus X protein induces transcriptional activation of a wide variety of viral and cellular genes. In addition to its ability to interact directly with many nuclear transcription factors, several reports indicate that the X protein stimulates different cytoplasmic kinase signal cascades. Using the yeast two-hybrid screen, we have isolated a clone designated X-associated protein 3 (XAP3) that encodes a human homolog of the rat protein kinase C-binding protein. One of the activation domains of X (amino acids 90 -122) is required for binding to XAP3, while the NH 2 -terminal part of XAP3 is necessary for binding to X. Both X and XAP3 bound specifically to the PKC isoenzyme synthesized in rabbit reticulocyte lysates. Overexpression of XAP3 enhanced X transactivation activity. These results support earlier findings that one of the mechanisms of transactivation by X is through involvement with the cellular protein kinase C pathway.Transcriptional activation is a widespread phenomenon among mammalian viral systems. Mammalian viral proteins that increase the rate of transcription can be divided into two groups based on whether they exhibit sequence-specific DNA binding. For example, the herpes simplex virus 1 (HSV-1) 1 Vmw175 (1, 2), the Epstein-Barr virus BZLF1 (3), the papilloma virus E2 (4), and the simian virus 40 and polyoma virus large T antigens (5) bind specific DNA sequence motifs, whereas HSV-1 VP16 (6, 7), the pseudorabies virus immediate early protein (8,9), and the adenovirus E1A protein (10, 11) do not. During the last decade, many studies have shown that non-DNA binding viral transactivators achieve their task by direct interaction with different cellular sequence-specific DNA binding transcription factors. For example, VP16 interacts with the Oct-1 protein, thereby positioning the VP16 activating domain at a promoter to enhance transcription (7,(12)(13)(14)(15)(16)(17). Similarly, E1A interacts with a number of cellular proteins including ATF-2 (18).The hepatitis B virus (HBV) X protein is a promiscuous transcriptional transactivator (reviewed in Ref. 19). This conclusion is derived from a large number of studies using mostly transient cotransfection of the bacterial chloramphenicol acetyltransferase (CAT) gene under control of a potential target promoter/enhancer and the X gene under the control of a heterologous promoter in mammalian cells. Induction of transcription by X usually ranges from 2-to 20-fold depending on the target promoter and cell type; whether this transactivation activity contributes to viral function, however, remains to be determined. Fusion of the X protein to the DNA-binding domain of the bacterial LexA repressor resulted in a protein that can activate transcription from a reporter plasmid bearing lexA operator sequences fused to a minimal promoter (20). Similarly, fusion of the X protein to the DNA-binding domain of transcription factor C/EBP increased the ability of X to activate a reporter containing C/EBP binding sites (21). Attempts to demonstrate sequence-speci...

Induction of the lactose-galactose regulon is strongly repressed by glucose in some but not all strains of Kluyveromyces lactis. We show here that in strongly repressed strains, two to three times less Kl-GAL4 mRNA is synthesized and that expression of structural genes in the regulon such as LAC4, the structural gene for 13-galactosidase lactose-galactose regulon. Strains such as Y1140 show weak repression because the fully induced level of the regulon, as measured by 0-galactosidase activity, is reduced only twofold by the presence of glucose in the culture medium (7). Strains such as CBS2360 show almost no repression (2), while strains such as JA6 (2) and Y1118 (this report) show strong (50-to 100-fold) repression of the regulon by glucose under inducing conditions. How might glucose affect expression of the regulon? Initial insight into the mechanism of glucose repression was provided by Breunig (2), who showed that a glucose-repressing strain could be converted to a nonrepressing strain by changing the allele carried at the K1-GAL4 locus (also called LAC9 [27]).Kl-GAL4 codes for a transcription activator protein that regulates and is essential for transcription of the lactosegalactose regulon (33). The Kl-GAL4 protein binds to a family of 17-bp DNA sequences, termed upstream activator sequences (UASG; the G refers to the lactose-galactose regulon), located in front of genes comprising the lactosegalactose regulon (3,9,17,25