Pre- and Post-Transcriptional Control of HBV Gene Expression: The Road Traveled towards the New Paradigm of HBx, Its Isoforms, and Their Diverse Functions.

Hepatitis B virus (HBV) is an enveloped DNA human virus belonging to the family. Perhaps its main distinguishable characteristic is the replication of its genome through a reverse transcription process. The HBV circular genome encodes only four overlapping reading frames, encoding for the main canonical proteins named core, P, surface, and X (or HBx protein).

Canonical and Divergent N-Terminal HBx Isoform Proteins Unveiled: Characteristics and Roles during HBV Replication.

Hepatitis B virus (HBV) X protein (HBx) is a viral regulatory and multifunctional protein. It is well-known that the canonical HBx reading frame bears two phylogenetically conserved internal in-frame translational initiation codons at Met2 and Met3, thus possibly generating divergent N-terminal smaller isoforms during translation. Here, we demonstrate that the three distinct HBx isoforms are generated from the ectopically expressed HBV HBx gene, named XF (full-length), XM (medium-length), and XS (short-length); they display different subcellular localizations when expressed individually in cultured hepatoma cells.

Phosphorylation of Phylogenetically Conserved Amino Acid Residues Confines HBx within Different Cell Compartments of Human Hepatocarcinoma Cells.

Hepatitis B virus (HBV) is a circular, and partially double-stranded DNA virus. Upon infection, the viral genome is translocated into the cell nucleus, generating the covalently closed circular DNA (cccDNA) intermediate, and forming a mini chromosome. HBV HBx is a small protein displaying multiple roles in HBV-infected cells, and in different subcellular locations.

The histone variant H3.3 regulates the transcription of the hepatitis B virus.

Introduction And Objectives: About 250 million people around the world are chronically infected with the hepatitis B virus (HBV). Those people are at risk of developing hepatocellular carcinoma. The HBV genome is organized as a minichromosome in the infected hepatocyte and is associated with histones and non-histone proteins.

Disrupting MLV integrase:BET protein interaction biases integration into quiescent chromatin and delays but does not eliminate tumor activation in a MYC/Runx2 mouse model.

Murine leukemia virus (MLV) integrase (IN) lacking the C-terminal tail peptide (TP) loses its interaction with the host bromodomain and extraterminal (BET) proteins and displays decreased integration at promoter/enhancers and transcriptional start sites/CpG islands. MLV lacking the IN TP via an altered open reading frame was used to infect tumorigenesis mouse model (MYC/Runx2) animals to observe integration patterns and phenotypic effects, but viral passage resulted in the restoration of the IN TP through small deletions. Mice subsequently infected with an MLV IN lacking the TP coding sequence (TP-) showed an improved median survival by 15 days compared to wild type (WT) MLV infection.

Recombinant HCV NS3 and NS5B enzymes exhibit multiple posttranslational modifications for potential regulation.

Posttranslational modification (PTM) of proteins is critical to modulate protein function and to improve the functional diversity of polypeptides. In this report, we have analyzed the PTM of both hepatitis C virus NS3 and NS5B enzyme proteins, upon their individual expression in insect cells under the baculovirus expression system. Using mass spectrometry, we present evidence that these recombinant proteins exhibit diverse covalent modifications on certain amino acid side chains, such as phosphorylation, ubiquitination, and acetylation.

The enzymes LSD1 and Set1A cooperate with the viral protein HBx to establish an active hepatitis B viral chromatin state.

With about 350 million people chronically infected around the world hepatitis B is a major health problem. Template for progeny HBV synthesis is the viral genome, organized as a minichromosome (cccDNA) inside the hepatocyte nucleus. How viral cccDNA gene expression is regulated by its chromatin structure; more importantly, how the modulation of this structure impacts on viral gene expression remains elusive.

Replication of a chronic hepatitis B virus genotype F1b construct.

Genotype F is one of the less-studied genotypes of human hepatitis B virus, although it is widely distributed in regions of Central and South American. Our previous studies have shown that HBV genotype F is prevalent in Chile, and phylogenetic analysis of its full-length sequence amplified from the sera of chronically infected patients identified it as HBV subgenotype F1b. We have previously reported the full-length sequence of a HBV molecular clone obtained from a patient chronically infected with genotype F1b.

Phosphorylation at the N-terminal finger subdomain of a viral RNA-dependent RNA polymerase.

The RNA-dependent RNA polymerase (RdRP) of the Hepatitis C virus (HCV), named NS5B, is phosphorylated by the cellular protein kinase C-related kinase 2 (PRK2) at two serine residues (Ser29 and Ser42) of the finger subdomain (genotype 1b). Herein, using bioinformatics, we selected four potential phosphorylation residues (Ser46, Ser76, Ser96 and Ser112) of NS5B (genotype 2a) for study. Whereas the NS5B Ser46D and Ser76D substitutions seemed to improve polymerase activity, the Ser96D mutation decreased colony formation efficiency.

Full-genome sequence of a hepatitis B virus genotype f1b clone from a chronically infected chilean patient.

The hepatitis B virus (HBV) is a DNA virus belonging to the Hepadnaviridae family. Viral isolates have been classified into 10 genotypes, named from A to J, and several subtypes. We report the full-genome sequence from a single molecular clone of HBV genotype F1b, amplified from a chronically infected Chilean patient.

Genotype F of hepatitis B: response to interferon.

Background: The relevance of HBV genotype diversity on interferon (IFN) therapy outcome in chronic hepatitis B patients has recently been highlighted. Data available for genotype F is poor. The aim of this work was to analyse the response of HBV genotype F to treatment with IFN.

Development of an enzyme-linked immunosorbent assay based on the murine leukemia virus p30 capsid protein.

Retroviral vectors derived from the murine leukemia virus (MuLV) are widely used as the starting material in the development of vectors for gene therapy and critical in answering questions relating to viral pathogenesis. The p30 capsid (CA) is the major viral core protein and an internal group antigen in MuLV. In this study, an enzyme-linked immunosorbent assay (ELISA) was developed for quantitation of MuLV infectious particles with p30 CA core antigen protein.

Genomic determinants of hepatitis C virus antiviral therapy outcomes: toward individualized treatment.

Hepatitis C virus (HCV) is an important global health problem with an estimated prevalence of more than 170 million infected individuals worldwide. Currently, the standard antiviral therapy, based on pegylated interferon alpha and ribavirin, can achieve a virological response in only nearly 50% of the patients infected with HCV genotype 1, the most widely distributed globally. During the last years, relevant data from genome-wide association studies (GWAS) about the impact and contribution of the patient genomics on viral infection outcomes has suggested the possibility that an individualized antiviral therapy can be considered.

The viral transactivator HBx protein exhibits a high potential for regulation via phosphorylation through an evolutionarily conserved mechanism.

Background: Hepatitis B virus (HBV) encodes an oncogenic factor, HBx, which is a multifunctional protein that can induce dysfunctional regulation of signaling pathways, transcription, and cell cycle progression, among other processes, through interactions with target host factors. The subcellular localization of HBx is both cytoplasmic and nuclear. This dynamic distribution of HBx could be essential to the multiple roles of the protein at different stages during HBV infection.

IL28B polymorphisms associated with therapy response in Chilean chronic hepatitis C patients.

Aim: To analyze the association of three IL28B single nucleotide polymorphisms with response to therapy in Chilean patients infected with hepatitis C virus (HCV).

Methods: We studied two groups of patients with chronic HCV infection (genotype 1), under standard combined treatment with pegylated interferon plus ribavirin. One group consisted of 50 patients with sustained virological response, whereas the second group consisted of 49 null responders.

Phylogenetic analysis of hepatitis B virus genotype F complete genome sequences from Chilean patients with chronic infection.

Molecular epidemiological data concerning the hepatitis B virus (HBV) in Chile are not known completely. Since the HBV genotype F is the most prevalent in the country, the goal of this study was to obtain full HBV genome sequences from patients infected chronically in order to determine their subgenotypes and the occurrence of resistance-associated mutations. Twenty-one serum samples from antiviral drug-naive patients with chronic hepatitis B were subjected to full-length PCR amplification, and both strands of the whole genomes were fully sequenced.

Inhibition of protease-inhibitor-resistant hepatitis C virus replicons and infectious virus by intracellular intrabodies.

Hepatitis C virus (HCV) infection is a common cause of chronic liver disease and a serious threat to human health. The HCV NS3/4A serine protease is necessary for viral replication and innate immune evasion, and represents a well-validated target for specific antiviral therapy. We previously reported the isolation of single-chain antibodies (scFvs) that inhibit NS3/4A protease activity in vitro.

miR-122 does not modulate the elongation phase of hepatitis C virus RNA synthesis in isolated replicase complexes.

miR-122 is an abundant, liver-specific microRNA that is required for efficient amplification of hepatitis C virus (HCV) RNA. Recent studies with a miR-122-specific locked nucleic acid antagomir have shown it to be an important host target for therapeutic intervention. However, considerable controversy exists concerning the mechanisms underlying the dependence of HCV replication on miR-122.

Molecular characterization of IPNV RNA replication intermediates during the viral infective cycle.

Infectious Pancreatic Necrosis Virus (IPNV) is a bisegmented, double-stranded RNA virus, which belongs to the Birnaviridae family. In the current study, we have analyzed the RNA replication intermediates (RI) purified throughout the viral replication cycle in cultured cells. Equilibrium ultracentrifugation of infected cellular lysates resulted in two major peaks of viral components.

Impaired replication of hepatitis C virus containing mutations in a conserved NS5B retinoblastoma protein-binding motif.

Hepatitis C virus (HCV) downregulates the retinoblastoma tumor suppressor protein (Rb), a central cell cycle regulator which is also targeted by oncoproteins expressed by DNA tumor viruses. HCV genome replication is also enhanced in proliferating cells. Thus, it is possible that HCV interactions with host cell cycle regulators, such as Rb, have evolved to modify the intracellular environment to promote viral replication.

Prevalence of hepatitis B virus genotypes in chronic carriers in Santiago, Chile.

The eight genotypes (designated A-H) of hepatitis B virus (HBV) display distinctive geographical distribution worldwide, with genotypes A, D and F frequently detected in South America. To determine the prevalence of HBV genotypes in Santiago, Chile, 131 samples from chronic carriers were used for PCR amplification, and genotyping was performed by RFLP. The results indicated that genotype F was the most prevalent among HBV carries (84% of the cases), whereas genotypes A, B, C and D were found at a prevalence of 3.

Production of infectious genotype 1a hepatitis C virus (Hutchinson strain) in cultured human hepatoma cells.

Infections with hepatitis C virus (HCV) are marked by frequent viral persistence, chronic liver disease, and extraordinary viral genetic diversity. Although much has been learned about HCV since its discovery, progress has been slowed by a lack of permissive cell culture systems supporting its replication. Productive infections have been achieved recently with genotype 2a virus, but cirrhosis and liver cancer are typically associated with genotype 1 HCV, which is more prevalent and relatively resistant to IFN therapy.

Interactions between virus proteins and host cell membranes during the viral life cycle.

The structure and function of cells are critically dependent on membranes, which not only separate the interior of the cell from its environment but also define the internal compartments. It is therefore not surprising that the major steps of the life cycle of viruses of animals and plants also depend on cellular membranes. Indeed, interactions of viral proteins with host cell membranes are important for viruses to enter into host cells, replicate their genome, and produce progeny particles.

Inhibition of virion-associated IPNV RNA polymerase, VP1, by radiolabeled nucleotide analogs.

Infectious pancreatic necrosis virus (IPNV) is a bi-segmented, dsRNA virus of the Birnaviridae family. The structural protein VP1 has been postulated as the RNA-dependent RNA polymerase (RdRp), but its transcriptional activity has not been unequivocally identified from viral particles. Here, we assayed partially purified IPNV in an in vitro RNA synthesis system.

Genome assembly and particle maturation of the birnavirus infectious pancreatic necrosis virus.

In this study, we have analyzed the morphogenesis of the birnavirus infectious pancreatic necrosis virus throughout the infective cycle in CHSE-214 cells by using a native agarose electrophoresis system. Two types of viral particles (designated A and B) were identified, isolated, and characterized both molecularly and biologically. Together, our results are consistent with a model of morphogenesis in which the genomic double-stranded RNA is immediately assembled, after synthesis, into a large (66-nm diameter) and uninfectious particle A, where the capsid is composed of both mature and immature viral polypeptides.