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Development of a Hepatitis B Virus Reporter System to Monitor the Early Stages of the Replication Cycle
Published on: February 1, 2017
[Hepatitis B virus replication and viral gene expressions do not affect CDC37 level in hepatocytes in vitro]
Chaowu Chen1, Bin Zhou, Ying Xu
1Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China.E-mail: ccwfimmu@126.com.
Objective:
To study the influence of hepatitis B virus (HBV) replication and expressions of different viral genes on CDC37 level in hepatocytes.
Methods:
We amplified and cloned 6 HBV genes (P, preS1, preS2, S, C and X) into pCMV expression vectors, which were transfected in Huh7 and HepG2 hepatoma cell lines, and CDC37 expression level in the cells was detected using Western blotting. Wealso cloned the promoter sequence of CDC37 into pGL3 vector, and co-transfected pGL3 with pCMV recombinant plasmids into Huh7 and HepG2 cells and the fluorescent signals were detected. To study the influence of HBV replication on CDC37 expression, we constructed 1.28-copy overlength genomes of HBV genotypes B, C, D and CD recombinant. The overlength HBV genomes were transformed into Adeasier-1 cells for recombination and into 293 cells for packaging. Huh7 and HepG2 cell lines infected with the packaged HBV recombinant adenoviruses were examined for CDC37 expression with Western blotting.
Results:
Western blotting showed that the expression of different HBV genes did not obviously affect the protein level of CDC37 in the hepatocytes. The protein expression of HBV genes had no effect on the activity of CDC37 promoter. Huh7 and HepG2 cells infected with 1.28-copy HBV replicon showed no significant changes in the expression level of CDC37.
Conclusion:
HBV replication and its gene expression have no effect on the level of CDC37 in hepatocytes in vitro.
