Requirement of CRTC1 coactivator for hepatitis B virus transcription

Nucleic Acids Research, Nov 2014

Transcription of hepatitis B virus (HBV) from the covalently closed circular DNA (cccDNA) template is essential for its replication. Suppressing the level and transcriptional activity of cccDNA might have anti-HBV effect. Although cellular transcription factors, such as CREB, which mediate HBV transcription, have been well described, transcriptional coactivators that facilitate this process are incompletely understood. In this study we showed that CREB-regulated transcriptional coactivator 1 (CRTC1) is required for HBV transcription and replication. The steady-state levels of CRTC1 protein were elevated in HBV-positive hepatoma cells and liver tissues. Ectopic expression of CRTC1 or its homolog CRTC2 or CRTC3 in hepatoma cells stimulated the activity of the preS2/S promoter of HBV, whereas overexpression of a dominant inactive form of CRTC1 inhibited HBV transcription. CRTC1 interacts with CREB and they are mutually required for the recruitment to the preS2/S promoter on cccDNA and for the activation of HBV transcription. Accumulation of pregenomic RNA (pgRNA) and cccDNA was observed when CRTC1 or its homologs were overexpressed, whereas the levels of pgRNA, cccDNA and secreted HBsAg were diminished when CRTC1 was compromised. In addition, HBV transactivator protein HBx stabilized CRTC1 and promoted its activity on HBV transcription. Our work reveals an essential role of CRTC1 coactivator in facilitating and supporting HBV transcription and replication.

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Requirement of CRTC1 coactivator for hepatitis B virus transcription

Hei-Man Vincent Tang 1 2 Wei-Wei Gao 1 2 Chi-Ping Chan 1 2 Yun Cheng 1 2 Vidyanath Chaudhary 1 2 Jian-Jun Deng 1 2 Kit-San Yuen 1 2 Chun-Ming Wong 0 1 Irene Oi-Lin Ng 0 1 Kin-Hang Kok 1 2 3 Jie Zhou 3 Dong-Yan Jin 1 2 0 Department of Pathology, The University of Hong Kong , Pokfulam , Hong Kong 1 State Key Laboratory for Liver Research, The University of Hong Kong , Pokfulam , Hong Kong 2 Department of Biochemistry, The University of Hong Kong , Pokfulam , Hong Kong 3 Department of Microbiology, The University of Hong Kong , Pokfulam , Hong Kong - Transcription of hepatitis B virus (HBV) from the covalently closed circular DNA (cccDNA) template is essential for its replication. Suppressing the level and transcriptional activity of cccDNA might have anti-HBV effect. Although cellular transcription factors, such as CREB, which mediate HBV transcription, have been well described, transcriptional coactivators that facilitate this process are incompletely understood. In this study we showed that CREBregulated transcriptional coactivator 1 (CRTC1) is required for HBV transcription and replication. The steady-state levels of CRTC1 protein were elevated in HBV-positive hepatoma cells and liver tissues. Ectopic expression of CRTC1 or its homolog CRTC2 or CRTC3 in hepatoma cells stimulated the activity of the preS2/S promoter of HBV, whereas overexpression of a dominant inactive form of CRTC1 inhibited HBV transcription. CRTC1 interacts with CREB and they are mutually required for the recruitment to the preS2/S promoter on cccDNA and for the activation of HBV transcription. Accumulation of pregenomic RNA (pgRNA) and cccDNA was observed when CRTC1 or its homologs were overexpressed, whereas the levels of pgRNA, cccDNA and secreted HBsAg were diminished when CRTC1 was compromised. In addition, HBV transactivator protein HBx stabilized CRTC1 and promoted its activity on HBV transcription. Our work reveals an essential role of CRTC1 coactivator in facilitating and supporting HBV transcription and replication. INTRODUCTION Hepatitis B virus (HBV), a member in the family of Hepadnaviridae, has a DNA genome 3.2 kb in size which is partially double stranded. Over 350 million people are chronically infected with HBV worldwide, including 10% of the population in Hong Kong and China, 1540% of whom will terminally develop severe liver diseases including hepatocellular carcinoma (HCC) (1). Although multiple factors are involved in the development of HCC in HBV carriers, high level of HBV DNA has been identified as a major risk (2). HBV infects hepatocytes that express its receptor named sodium taurocholate cotransporting polypeptide (3). Upon viral entry and release of genetic material, covalently closed circular DNA (cccDNA) is generated from relaxed circular DNA (rcDNA) and complexed with different viral proteins, histones, transcription factors and coactivators to form a nuclear minichromosome, which serves as a central template for all HBV transcription (4,5). Transcription of pregenomic RNA (pgRNA) from cccDNA is rate-limiting in genome amplification and replication. cccDNA can be amplified through an unknown mechanism (6). The stability of the cccDNA pool is a major determinant in viral clearance. cccDNA is refractory to antivirals such as nucleoside or nucleotide analogs. cccDNA is also accounted for viral relapse after cessation of anti-HBV therapy (5). cccDNA is therefore an important target for better control and elimination of HBV infection (7,8). In line with this, understanding the mechanism by which HBV transcription from cccDNA is regulated might reveal new strategies for therapeutic intervention. HBV transcription is mediated by transcription factors recruited to cccDNA including CREB, ATF1, STAT1 and KLF15 (4,9). CREB is essential for HBV replication and it binds to the cAMP response elements (CREs) located at the X and preS2 promoters of the viral genome (10,11). Transcriptional coactivators such as p300, pCAF and CREB *To whom correspondence should be addressed. Tel: +852 3917 9491; Fax: +852 2855 1254; Email: The authors wish it to be known that, in their opinion, the first three authors should be regarded as Joint First Authors. binding protein (CBP) are also known to be recruited to cccDNA to support HBV transcription through acetylation of histones and transcription factors (12). In addition, CREB-regulated transcriptional coactivator 2 (CRTC2) is also thought to potentiate HBV transcription through the induction of PGC1 (13). However, the roles of additional CRTC transcriptional coactivators in relation to the function of CREB in HBV transcription have not been characterized. CRTCs are potent coactivators of CREB-dependent transcription from both canonical cellular CREs and the non-canonical viral CREs found in the long terminal repeats of the oncogenic retrovirus human T-cell leukemia virus type 1 (14,15). There are three isoforms of human CRTCs, namely CRTC1, CRTC2 and CRTC3. They interact with CR (...truncated)


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Hei-Man Vincent Tang, Wei-Wei Gao, Chi-Ping Chan, Yun Cheng, Vidyanath Chaudhary, Jian-Jun Deng, Kit-San Yuen, Chun-Ming Wong, Irene Oi-Lin Ng, Kin-Hang Kok, Jie Zhou, Dong-Yan Jin. Requirement of CRTC1 coactivator for hepatitis B virus transcription, Nucleic Acids Research, 2014, pp. 12455-12468, 42/20, DOI: 10.1093/nar/gku925