Investigation of Protein-DNA Interactions
in Enhancer Ⅱand Core Promoter of HBV by in vivo Footprinting
ZHAO Xin-Tai�L, YOU Zhong-Sheng, CHENG Ping and
WANG Yuan*
( Shanghai Institute of Biochemistry, the Chinese Academy of Sciences, Shanghai
200031, China )
Abstract The protein-DNA interactions
in the enhancer Ⅱ (ENⅡ) and core promoter regions of hepatitis B virus (HBV)
were investigated by the ligation-mediated polymerase chain reaction (LMPCR) in
vivo footprinting in HepG2.2.15 cell line. Major footprints are evident at
the enhancer Ⅱ and core promoter regions. In particular, proteins appear to
interact with the sites for Sp1, C/EBP, HNF4, HNF3, HNF1 and hB1F, as well as
the TATA-like element. It confirmed that these protein-DNA interactions defined
in vitro play important roles in the regulation of 3.5 kb RNAs
transcription of HBV in vivo. Notably, three novel sites nt 1774-1789,
nt 1801-1818 and nt 1835-1843 were found to be protected or hypersensitive,
indicating that they were also involved in protein-DNA interactions in vivo.
Functional analysis using CAT reporter gene demonstrated that the site
from nt 1801 to 1818 was important for the function of HBV core promoter.
Key Words hepatitis B virus (HBV)； LMPCR in vivo
footprinting； enhancer Ⅱ (ENⅡ)； core promoter； CAT assay
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