Title: Hepatitis B virus (HBV) particles a system by transient expression of t (DNA transfection/hepatitis B virus replication/pregenome RNA/in
Abstract: An in vitro system for the production of hepatitis B virus (HBV) particles was established by the transient expression of transfected HBV DNA using a human hepatocellular carcinoma cell, HuH-7, as a recipient. The 3.6- and 2.2-kilobase transcripts observed were similar to those in virus-infected liver cells. Both transcripts revealed the microheterogeneity of their 5' ends. The formation of virus- related particles subsequent to the RNA transcription was demonstrated. The core particles observed in the cytoplasm and the virus particles secreted in the culture medium con- tained the replicative intermediates of IBV DNA and banded at densities of 1.35-1.36 g/cm3 and 1.22-1.24 g/cm3, respec- tively. Furthermore, the in vitro mutagenesis of the template HBV DNA demonstrated that the P gene as well as the C gene products were essential for the production of HBV particles. Infection of hepatitis B virus (HBV) causes acute and chronic hepatitis. Furthermore, HBV is known to be a major cause of human liver cancer. HBV DNA has the following unique structural characteristics (1). Two linear DNA strands (plus and minus) form a circular molecule by annealing each cohesive end region. The minus strand is 3.2 kilobases (kb) in size with a protein linked to its 5' end. The plus strand usually shows various degrees of incompleteness. Similar HBV-like animal viruses have been discovered in wood- chucks (woodchuck hepatitis virus), ground squirrels (ground squirrel hepatitis virus, GSHV), and ducks (duck hepatitis B virus, DHBV). The replication cycle of the virus was proposed by Summers and Mason based on their observations of the in vivo replication of DHBV (2). How- ever, the restricted host range of HBV and lack of a cell culture system for virus production have been major prob- lems hindering from understanding of HBV replication and transcription at the molecular level. In this investigation, an in vitro system for the production of HBV particles was established by the transient expression of transfected HBV DNA, using a human hepatocellular carcinoma cell as a recipient. The RNA transcription of HBV DNA, similar to that in virus infection in vivo, was observed. Furthermore, mutagenesis of the template HBV DNA indi- cated virus production to be absolutely dependent on the template plasmid and the P gene as well as the C gene products to be essential for the synthesis of HBV DNA.
Publication Year: 2016
Publication Date: 2016-01-01
Language: en
Type: article
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