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)