News and Publications

Inhibition of Hepatitis B Virus Transcription and Replication by Hepatocyte Nuclear Factor 3b

H. Tang, K.E. Banks, A.L. Anderson, A. McLachlan

Infection with hepatitis B virus (HBV), which causes both acute and chronic disease, is a worldwide health problem. No reliable treatment exists for the estimated 200 million to 500 million persons who are chronic HBV carriers. Therefore, antiviral strategies are urgently needed to control the liver disease associated with HBV infection.

HBV replicates essentially exclusively in the liver by reverse transcription of the 3.5-kb pregenomic RNA, generating the partially double-stranded 3.2-kb HBV DNA genome. Viral tropism is probably determined, in part, by a liver-specific receptor that is required for HBV infection of hepatocytes. However, viral replication is also restricted to hepatocytes because of the essential requirement for the liver-enriched transcription factors hepatocyte nuclear factor (HNF) 4 and retinoid X receptor a-peroxisome proliferator activated receptor a that regulate the synthesis of the pregenomic RNA. The level of synthesis of the pregenomic RNA is governed by the activity of the nucleocapsid promoter. A variety of ubiquitous and liver-enriched transcription factors in addition to the nuclear hormone receptors, HNF4, and retinoid X receptor a-peroxisome proliferator activated receptor a appear to modulate the rate of initiation of transcription from the nucleocapsid promoter. However, nuclear hormone receptor-dependent viral replication can be inhibited by the liver-enriched transcription factor HNF3 because of the preferential inhibition of 3.5-kb pregenomic RNA synthesis relative to 3.5-kb precore RNA synthesis.

We investigated the mechanism of regulation of HBV 3.5-kb RNA synthesis and inhibition of viral replication by HNF3. We examined the functional domain of HNF3b responsible for regulating nuclear hormone receptor-dependent HBV 3.5-kb RNA synthesis and viral replication. We found that the amino-terminal transcriptional activation domain of HNF3b was primarily responsible for inhibiting viral replication, whereas the carboxyl-terminal transcriptional activation domain did not greatly affect HBV DNA synthesis. The inhibitory effect of the former HNF3b domain on viral replication contrasts with the observation that the amino-terminal transcriptional activation domain was responsible for the increase in the expression of reporter genes mediated by the HBV large surface antigen and nucleocapsid promoters.

These results suggested that the lower level of the HBV 3.5-kb RNA might occur because HNF3b reduced the rate of HBV 3.5-kb RNA elongation rather than negatively regulating the activity of the nucleocapsid promoter. This possibility was supported by the observation that HNF3b could reduce viral replication when pregenomic RNA was synthesized from the cytomegalovirus immediate-early promoter rather than from the HBV nucleocapsid promoter. In addition, the ability of HNF3b to preferentially decrease the level of the pregenomic RNA compared with precore RNA produced conditions in which the precore RNA-encoded hepatitis B e antigen also contributed to the reduction in viral biosynthesis. Therefore, HNF3b apparently inhibits HBV replication by reducing pregenomic RNA abundance by transcriptional interference and modulating the effect of hepatitis B e antigen on viral biosynthesis.

We used a transgenic mouse model of chronic HBV infection to examine the role of HNF3b in the regulation of viral transcription and replication in vivo. HBV transgenic mice were bred with transgenic mice that overexpress HNF3b in the liver. Doubly transgenic mice with elevated levels of HNF3b expression were examined for viral transcription and replication. The results indicated that HNF3b inhibits HBV replication in the liver in a manner similar to that observed in cell culture. Viral replication was reduced by HNF3b overexpression to a significantly greater extent than was the level of the HBV 3.5-kb RNA. This finding suggests that physiologic or therapeutic stimuli that modulate HBV 3.5-kb RNA synthesis to a modest extent may have marked antiviral potential.

PUBLICATIONS

Alcantara, F., Tang, H., McLachlan, A. Functional characterization of the interferon regulatory element in the enhancer 1 region of the hepatitis B virus genome. Nucleic Acids Res. 30:2068, 2002.

Banks, K.E., Anderson, A.L., Tang, H., Hughes, D.E., Costa, R.H., McLachlan, A. Hepatocyte nuclear factor 3b inhibits hepatitis B virus replication in vivo. J. Virol. 76:12974, 2002.

Tang, H., Banks, K.E., Anderson, A.L., McLachlan, A. Hepatitis B virus transcription and replication. Drug News Perspect. 14:325, 2001.

Tang, H., McLachlan, A. Avian and mammalian hepadnaviruses have distinct transcription factor requirements for viral replication. J. Virol. 76:7468, 2002.

Tang, H., McLachlan, A. Mechanisms of inhibition of nuclear hormone receptor dependent hepatitis B virus replication by hepatocyte nuclear factor 3b. J. Virol. 76:8572, 2002.

Tang, H., McLachlan, A. A pregenomic RNA sequence adjacent to DRI and complementary to epsilon determines hepatitis B virus replicaton efficiency. Virology, in press.