In Vitro Replication Models for the Hepatitis C Virus
In Vitro Replication Models for the Hepatitis C Virus
Soon after the discovery of the hepatitis C virus (HCV), attention turned to the development of models whereby replication of the virus could be investigated. Among the HCV replication models developed, the HCV RNA replicon model and the newly discovered infectious cell culture systems have had an immediate impact on the study of HCV replication, and will continue to lead to important advances in our understanding of HCV replication. The aim of this study is to deal with developments in HCV replication models in a chronological order from the early 1990s to the recent infectious HCV cell culture systems.
The hepatitis C virus (HCV) was discovered in 1989, and infects 170 million people. HCV is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. HCV is a positive strand RNA virus and forms the sole member of a separate genus, Hepacivirus, within the Flaviviridae virus family. The HCV genome is composed of a large open reading frame (ORF) flanked by 5'- and 3'-untranslated regions (UTR). The ORF is processed into structural (core, E1 and E2) and nonstructural (NS) proteins (NS2, NS3, NS4a, NS4b, NS5a and NS5b). (Fig. 1). The synthesis of a protein encoded by an alternative reading frame within the core region has been reported by several groups. It is designated as alternative reading frame protein or frameshift protein and is dispensable for HCV RNA replication, but whether it is expressed during natural HCV infection is unknown. In addition, p7 is a 63 amino acid polypeptide located at the junction between the structural region and the NS region. p7 is composed of two transmembrane domains and has recently been reported to be involved in ion channel activity. By sequence comparison at the 5'-UTR and NS5b regions, HCV isolates are placed in one of six genotypes and various subtypes. In addition, HCV sequence variability is observed within a single HCV infected individual (termed quasispecies). HCV diversity is generally attributed to the (i) error-prone RNA-directed RNA polymerase (NS5b), (ii) high rate of HCV replication and (iii) long-term persistent HCV infection.
(Enlarge Image)
The genomic organization of the HCV. Core, core protein; ARFP, alternate reading frame protein; E1, envelope 1 glycoprotein; E2, envelope 2 glycoprotein; NS2, nonstructural protein 2; NS3, nonstructural protein 3; 4A, nonstructural protein 4A; 4B, nonstructural protein 4B; NS5A, nonstructural protein 5A; NS5B, nonstructural protein 5B; 3'-UTR, 3'-untranslated region; 5'-UTR, 5'-untranslated region; +ssRNA, positive single strand RNA genome.
Numerous in vitro models for the investigation of HCV replication have been developed. However, until very recently these pursuits did not yield a robust HCV replication model in which the complete life cycle of HCV could be studied by direct biochemical means. The basis of this review is to discuss the principal in vitro replication models developed for the HCV (Fig. 2). These models will incorporate the development of engineered virus strains, mammalian systems expressing HCV in a variety of mechanisms, RNA replicons and finally the recently described robust in vitro models for hepatitis C virion production.
(Enlarge Image)
The development of in vitro replication models from the discovery of hepatitis C virus (1989) to the recently described model reporting the production of infectious hepatitis C virions in cell culture (2005). A number of in vitro models were constructed and investigated throughout this time period and are discussed in this review. HCV, hepatitis C virus.
Soon after the discovery of the hepatitis C virus (HCV), attention turned to the development of models whereby replication of the virus could be investigated. Among the HCV replication models developed, the HCV RNA replicon model and the newly discovered infectious cell culture systems have had an immediate impact on the study of HCV replication, and will continue to lead to important advances in our understanding of HCV replication. The aim of this study is to deal with developments in HCV replication models in a chronological order from the early 1990s to the recent infectious HCV cell culture systems.
The hepatitis C virus (HCV) was discovered in 1989, and infects 170 million people. HCV is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. HCV is a positive strand RNA virus and forms the sole member of a separate genus, Hepacivirus, within the Flaviviridae virus family. The HCV genome is composed of a large open reading frame (ORF) flanked by 5'- and 3'-untranslated regions (UTR). The ORF is processed into structural (core, E1 and E2) and nonstructural (NS) proteins (NS2, NS3, NS4a, NS4b, NS5a and NS5b). (Fig. 1). The synthesis of a protein encoded by an alternative reading frame within the core region has been reported by several groups. It is designated as alternative reading frame protein or frameshift protein and is dispensable for HCV RNA replication, but whether it is expressed during natural HCV infection is unknown. In addition, p7 is a 63 amino acid polypeptide located at the junction between the structural region and the NS region. p7 is composed of two transmembrane domains and has recently been reported to be involved in ion channel activity. By sequence comparison at the 5'-UTR and NS5b regions, HCV isolates are placed in one of six genotypes and various subtypes. In addition, HCV sequence variability is observed within a single HCV infected individual (termed quasispecies). HCV diversity is generally attributed to the (i) error-prone RNA-directed RNA polymerase (NS5b), (ii) high rate of HCV replication and (iii) long-term persistent HCV infection.
(Enlarge Image)
The genomic organization of the HCV. Core, core protein; ARFP, alternate reading frame protein; E1, envelope 1 glycoprotein; E2, envelope 2 glycoprotein; NS2, nonstructural protein 2; NS3, nonstructural protein 3; 4A, nonstructural protein 4A; 4B, nonstructural protein 4B; NS5A, nonstructural protein 5A; NS5B, nonstructural protein 5B; 3'-UTR, 3'-untranslated region; 5'-UTR, 5'-untranslated region; +ssRNA, positive single strand RNA genome.
Numerous in vitro models for the investigation of HCV replication have been developed. However, until very recently these pursuits did not yield a robust HCV replication model in which the complete life cycle of HCV could be studied by direct biochemical means. The basis of this review is to discuss the principal in vitro replication models developed for the HCV (Fig. 2). These models will incorporate the development of engineered virus strains, mammalian systems expressing HCV in a variety of mechanisms, RNA replicons and finally the recently described robust in vitro models for hepatitis C virion production.
(Enlarge Image)
The development of in vitro replication models from the discovery of hepatitis C virus (1989) to the recently described model reporting the production of infectious hepatitis C virions in cell culture (2005). A number of in vitro models were constructed and investigated throughout this time period and are discussed in this review. HCV, hepatitis C virus.
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