Hepatitis B viral polymerase fusion proteins are biologically active and can interact with the hepatitis C virus core protein in vivo

K. L. Chen, C. M. Chen, C. M. Shih, H. L. Huang, Y. H W Lee, C. Chang, S. J. Lo

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Hepadnaviruses and retroviruses are evolutionarily related families because they both require a process of reverse transcription for genome replication. However, hepadnaviruses produce polymerase (pol) and core proteins separately, while retroviruses synthesize a gag-pol fusion protein that is subsequently cleaved by a virally encoded protease to release a functional polymerase. To test whether an additional sequence at the N-terminus of pol in hepatitis B virus (HBV) interferes with its function, we created two plasmids expressing core-pol fusion proteins, core144-pol and core31-pol. Secreted particles obtained from HuH-7 cells, which were cotransfected with a core-pol fusion protein-expressing plasmid and a core-expressing plasmid, showed a positive signal of HBV DNA by the endogenous polymerase assay, indicating that the core-pol fusion proteins retain DNA priming, polymerization and RNase H activities. The fusion protein was detected in the cytoplasm of transfected cells and in secreted virions by immunoprecipitation. Furthermore, we found by immunofluorescence staining that the HBV core-pol fusion protein colocalized with the hepatitis C virus (HCV) core protein in cytoplasm and in lipid droplets. Immunoprecipitation studies showed that the anti-HCV core complex contained the HBV core-pol fusion protein while the anti-HBV pol complex contained the HCV core protein, which supports the hypothesis that the HCV core protein can form a complex with the HBV core-pol fusion protein.

Original languageEnglish
Pages (from-to)492-503
Number of pages12
JournalJournal of Biomedical Science
Volume8
Issue number6
DOIs
Publication statusPublished - 2001
Externally publishedYes

Fingerprint

Viral Fusion Proteins
Hepatitis B
Fusion reactions
Viruses
Hepatitis B virus
Proteins
Hepadnaviridae
Plasmids
Retroviridae
Immunoprecipitation
Cytoplasm
Ribonuclease H
Hepatitis C virus nucleocapsid protein
DNA
DNA-Directed DNA Polymerase
Transcription
Polymerization
Hepacivirus
Virion
Reverse Transcription

Keywords

  • Endogenous polymerase activity
  • Evolutionary distance
  • Hepatitis C virus core protein
  • Immunofluorescence microscopy
  • Pol localization
  • Reverse transcription

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Hepatitis B viral polymerase fusion proteins are biologically active and can interact with the hepatitis C virus core protein in vivo. / Chen, K. L.; Chen, C. M.; Shih, C. M.; Huang, H. L.; Lee, Y. H W; Chang, C.; Lo, S. J.

In: Journal of Biomedical Science, Vol. 8, No. 6, 2001, p. 492-503.

Research output: Contribution to journalArticle

Chen, K. L. ; Chen, C. M. ; Shih, C. M. ; Huang, H. L. ; Lee, Y. H W ; Chang, C. ; Lo, S. J. / Hepatitis B viral polymerase fusion proteins are biologically active and can interact with the hepatitis C virus core protein in vivo. In: Journal of Biomedical Science. 2001 ; Vol. 8, No. 6. pp. 492-503.
@article{cf2a5ccb6dd5411098ca1b9bd91a2aa8,
title = "Hepatitis B viral polymerase fusion proteins are biologically active and can interact with the hepatitis C virus core protein in vivo",
abstract = "Hepadnaviruses and retroviruses are evolutionarily related families because they both require a process of reverse transcription for genome replication. However, hepadnaviruses produce polymerase (pol) and core proteins separately, while retroviruses synthesize a gag-pol fusion protein that is subsequently cleaved by a virally encoded protease to release a functional polymerase. To test whether an additional sequence at the N-terminus of pol in hepatitis B virus (HBV) interferes with its function, we created two plasmids expressing core-pol fusion proteins, core144-pol and core31-pol. Secreted particles obtained from HuH-7 cells, which were cotransfected with a core-pol fusion protein-expressing plasmid and a core-expressing plasmid, showed a positive signal of HBV DNA by the endogenous polymerase assay, indicating that the core-pol fusion proteins retain DNA priming, polymerization and RNase H activities. The fusion protein was detected in the cytoplasm of transfected cells and in secreted virions by immunoprecipitation. Furthermore, we found by immunofluorescence staining that the HBV core-pol fusion protein colocalized with the hepatitis C virus (HCV) core protein in cytoplasm and in lipid droplets. Immunoprecipitation studies showed that the anti-HCV core complex contained the HBV core-pol fusion protein while the anti-HBV pol complex contained the HCV core protein, which supports the hypothesis that the HCV core protein can form a complex with the HBV core-pol fusion protein.",
keywords = "Endogenous polymerase activity, Evolutionary distance, Hepatitis C virus core protein, Immunofluorescence microscopy, Pol localization, Reverse transcription",
author = "Chen, {K. L.} and Chen, {C. M.} and Shih, {C. M.} and Huang, {H. L.} and Lee, {Y. H W} and C. Chang and Lo, {S. J.}",
year = "2001",
doi = "10.1159/000046171",
language = "English",
volume = "8",
pages = "492--503",
journal = "Journal of Biomedical Science",
issn = "1021-7770",
publisher = "BioMed Central",
number = "6",

}

TY - JOUR

T1 - Hepatitis B viral polymerase fusion proteins are biologically active and can interact with the hepatitis C virus core protein in vivo

AU - Chen, K. L.

AU - Chen, C. M.

AU - Shih, C. M.

AU - Huang, H. L.

AU - Lee, Y. H W

AU - Chang, C.

AU - Lo, S. J.

PY - 2001

Y1 - 2001

N2 - Hepadnaviruses and retroviruses are evolutionarily related families because they both require a process of reverse transcription for genome replication. However, hepadnaviruses produce polymerase (pol) and core proteins separately, while retroviruses synthesize a gag-pol fusion protein that is subsequently cleaved by a virally encoded protease to release a functional polymerase. To test whether an additional sequence at the N-terminus of pol in hepatitis B virus (HBV) interferes with its function, we created two plasmids expressing core-pol fusion proteins, core144-pol and core31-pol. Secreted particles obtained from HuH-7 cells, which were cotransfected with a core-pol fusion protein-expressing plasmid and a core-expressing plasmid, showed a positive signal of HBV DNA by the endogenous polymerase assay, indicating that the core-pol fusion proteins retain DNA priming, polymerization and RNase H activities. The fusion protein was detected in the cytoplasm of transfected cells and in secreted virions by immunoprecipitation. Furthermore, we found by immunofluorescence staining that the HBV core-pol fusion protein colocalized with the hepatitis C virus (HCV) core protein in cytoplasm and in lipid droplets. Immunoprecipitation studies showed that the anti-HCV core complex contained the HBV core-pol fusion protein while the anti-HBV pol complex contained the HCV core protein, which supports the hypothesis that the HCV core protein can form a complex with the HBV core-pol fusion protein.

AB - Hepadnaviruses and retroviruses are evolutionarily related families because they both require a process of reverse transcription for genome replication. However, hepadnaviruses produce polymerase (pol) and core proteins separately, while retroviruses synthesize a gag-pol fusion protein that is subsequently cleaved by a virally encoded protease to release a functional polymerase. To test whether an additional sequence at the N-terminus of pol in hepatitis B virus (HBV) interferes with its function, we created two plasmids expressing core-pol fusion proteins, core144-pol and core31-pol. Secreted particles obtained from HuH-7 cells, which were cotransfected with a core-pol fusion protein-expressing plasmid and a core-expressing plasmid, showed a positive signal of HBV DNA by the endogenous polymerase assay, indicating that the core-pol fusion proteins retain DNA priming, polymerization and RNase H activities. The fusion protein was detected in the cytoplasm of transfected cells and in secreted virions by immunoprecipitation. Furthermore, we found by immunofluorescence staining that the HBV core-pol fusion protein colocalized with the hepatitis C virus (HCV) core protein in cytoplasm and in lipid droplets. Immunoprecipitation studies showed that the anti-HCV core complex contained the HBV core-pol fusion protein while the anti-HBV pol complex contained the HCV core protein, which supports the hypothesis that the HCV core protein can form a complex with the HBV core-pol fusion protein.

KW - Endogenous polymerase activity

KW - Evolutionary distance

KW - Hepatitis C virus core protein

KW - Immunofluorescence microscopy

KW - Pol localization

KW - Reverse transcription

UR - http://www.scopus.com/inward/record.url?scp=0034766897&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034766897&partnerID=8YFLogxK

U2 - 10.1159/000046171

DO - 10.1159/000046171

M3 - Article

C2 - 11702013

AN - SCOPUS:0034766897

VL - 8

SP - 492

EP - 503

JO - Journal of Biomedical Science

JF - Journal of Biomedical Science

SN - 1021-7770

IS - 6

ER -