scholarly journals RNA-binding activity of hepatitis delta antigen involves two arginine-rich motifs and is required for hepatitis delta virus RNA replication.

1993 ◽  
Vol 67 (4) ◽  
pp. 2221-2227 ◽  
Author(s):  
C Z Lee ◽  
J H Lin ◽  
M Chao ◽  
K McKnight ◽  
M M Lai
Keyword(s):  
Hepatitis Delta Virus ◽  
Rna Binding ◽  
Rna Replication ◽  
Binding Activity ◽  
Hepatitis Delta ◽  
Delta Antigen ◽  
Virus Rna ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

scholarly journals Arginine-Rich Motifs Are Not Required for Hepatitis Delta Virus RNA Binding Activity of the Hepatitis Delta Antigen

2013 ◽  
Vol 87 (15) ◽  
pp. 8665-8674 ◽  
Author(s):  
L. H. Daigh ◽  
B. L. Griffin ◽  
A. Soroush ◽  
M. R. Mamedov ◽  
J. L. Casey
Keyword(s):  
Hepatitis Delta Virus ◽  
Rna Binding ◽  
Binding Activity ◽  
Hepatitis Delta ◽  
Delta Antigen ◽  
Virus Rna ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

scholarly journals Large Hepatitis Delta Antigen Is Not a Suppressor of Hepatitis Delta Virus RNA Synthesis once RNA Replication Is Established

2002 ◽  
Vol 76 (19) ◽  
pp. 9910-9919 ◽  
Author(s):  
Thomas B. Macnaughton ◽  
Michael M. C. Lai
Keyword(s):  
Hepatitis Delta Virus ◽  
Rna Synthesis ◽  
State Level ◽  
Rna Replication ◽  
Replication Cycle ◽  
Steady State Level ◽  
Hepatitis Delta ◽  
Delta Antigen ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

ABSTRACT Moderation of hepatitis delta virus (HDV) replication is a likely prerequisite in the establishment of chronic infections and is thought to be mediated by the intracellular accumulation of large hepatitis delta antigen (L-HDAg). The regulatory role of this protein was suggested from several studies showing that cotransfection of plasmid cDNAs expressing both L-HDAg and HDV RNA results in a potent inhibition of HDV RNA replication. However, since this approach differs significantly from natural HDV infections, where HDV RNA replication is initiated from an RNA template, and L-HDAg appears only late in the replication cycle, it remains unclear whether L-HDAg can modulate HDV RNA replication in the natural HDV replication cycle. In this study, we investigated the effect of L-HDAg, produced as a result of the natural HDV RNA editing event, on HDV RNA replication. The results showed that following cDNA-free HDV RNA transfection, a steady-state level of RNA was established at 3 to 4 days posttransfection. The same level of HDV RNA was reached when a mutant HDV genome unable to make L-HDAg was used, suggesting that L-HDAg did not play a role. The rates of HDV RNA synthesis, as measured by metabolic labeling experiments, were identical at 4 and 8 days posttransfection and in the wild type and the L-HDAg-deficient mutant. We further examined the effect of overexpression of L-HDAg at various stages of the HDV replication cycle, showing that HDV RNA synthesis was resistant to L-HDAg when it was overexpressed 3 days after HDV RNA replication had initiated. Finally, we showed that, contrary to conventional thinking, L-HDAg alone, at a certain molar ratio with HDV RNA, can initiate HDV RNA replication. Thus, L-HDAg does not inherently inhibit HDV RNA synthesis. Taken together, these results indicated that L-HDAg affects neither the rate of HDV RNA synthesis nor the final steady-state level of HDV RNA and that L-HDAg is unlikely to act as an inhibitor of HDV RNA replication in the natural HDV replication cycle.

scholarly journals Hepatitis Delta Virus RNA Editing Is Highly Specific for the Amber/W Site and Is Suppressed by Hepatitis Delta Antigen

1998 ◽  
Vol 18 (4) ◽  
pp. 1919-1926 ◽  
Author(s):  
Andrew G. Polson ◽  
Herbert L. Ley ◽  
Brenda L. Bass ◽  
John L. Casey
Keyword(s):  
Rna Editing ◽  
Hepatitis Delta Virus ◽  
Hepatitis Delta ◽  
Reading Frame ◽  
Adenosine Deaminases ◽  
Delta Antigen ◽  
Virus Rna ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

ABSTRACT RNA editing at adenosine 1012 (amber/W site) in the antigenomic RNA of hepatitis delta virus (HDV) allows two essential forms of the viral protein, hepatitis delta antigen (HDAg), to be synthesized from a single open reading frame. Editing at the amber/W site is thought to be catalyzed by one of the cellular enzymes known as adenosine deaminases that act on RNA (ADARs). In vitro, the enzymes ADAR1 and ADAR2 deaminate adenosines within many different sequences of base-paired RNA. Since promiscuous deamination could compromise the viability of HDV, we wondered if additional deamination events occurred within the highly base paired HDV RNA. By sequencing cDNAs derived from HDV RNA from transfected Huh-7 cells, we determined that the RNA was not extensively modified at other adenosines. Approximately 0.16 to 0.32 adenosines were modified per antigenome during 6 to 13 days posttransfection. Interestingly, all observed non-amber/W adenosine modifications, which occurred mostly at positions that are highly conserved among naturally occurring HDV isolates, were found in RNAs that were also modified at the amber/W site. Such coordinate modification likely limits potential deleterious effects of promiscuous editing. Neither viral replication nor HDAg was required for the highly specific editing observed in cells. However, HDAg was found to suppress editing at the amber/W site when expressed at levels similar to those found during HDV replication. These data suggest HDAg may regulate amber/W site editing during virus replication.

scholarly journals Hepatitis Delta Antigen Mediates the Nuclear Import of Hepatitis Delta Virus RNA

1998 ◽  
Vol 72 (5) ◽  
pp. 3684-3690 ◽  
Author(s):  
Huei-Chi Chou ◽  
Tsai-Yuan Hsieh ◽  
Gwo-Tarng Sheu ◽  
Michael M. C. Lai
Keyword(s):  
Nuclear Import ◽  
Hepatitis Delta Virus ◽  
Rna Binding ◽  
Binding Motif ◽  
Hepatitis Delta ◽  
Binding Motifs ◽  
Delta Antigen ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

ABSTRACT Hepatitis delta virus (HDV) RNA replicates in the nuclei of virus-infected cells. The mechanism of nuclear import of HDV RNA is so far unknown. Using a fluorescein-labeled HDV RNA introduced into partially permeabilized HeLa cells, we found that HDV RNA accumulated only in the cytoplasm. However, in the presence of hepatitis delta antigen (HDAg), which is the only protein encoded by HDV RNA, the HDV RNA was translocated into the nucleus, suggesting that nuclear import of HDV RNA is mediated by HDAg. Deletion of the nuclear localization signal (NLS) or RNA-binding motifs of HDAg resulted in the failure of nuclear import of HDV RNA, indicating that both the NLS and an RNA-binding motif of HDAg are required for the RNA-transporting activity of HDAg. Surprisingly, any one of the three previously identified RNA-binding motifs was sufficient to confer the RNA-transporting activity. We have further shown that HDAg, via its NLS, interacts with karyopherin α2 in vitro, suggesting that nuclear import of the HDAg-HDV RNA complex is mediated by the karyopherin α2β heterodimer. The nuclear import of HDV RNA may be the first biological function of HDAg in the HDV life cycle.

scholarly journals Genotype-Specific Complementation of Hepatitis Delta Virus RNA Replication by Hepatitis Delta Antigen

1998 ◽  
Vol 72 (4) ◽  
pp. 2806-2814 ◽  
Author(s):  
John L. Casey ◽  
John L. Gerin
Keyword(s):  
Hepatitis Delta Virus ◽  
Rna Replication ◽  
Cdna Clones ◽  
Genotype I ◽  
Hepatitis Delta ◽  
Genotype Iii ◽  
Delta Antigen ◽  
Hepatitis Delta Antigen ◽  
Hdv Genotype ◽  
Delta Virus

ABSTRACT Characterizations of genetic variations among hepatitis delta virus (HDV) isolates have focused principally on phylogenetic analysis of sequences, which vary by 30 to 40% among three genotypes and about 10 to 15% among isolates of the same genotype. The significance of the sequence differences has been unclear but could be responsible for pathogenic variations associated with the different genotypes. Studies of the mechanisms of HDV replication have been limited to cDNA clones from HDV genotype I, which is the most common. To perform a comparative analysis of HDV RNA replication in genotypes I and III, we have obtained a full-length cDNA clone from an HDV genotype III isolate. In transfected Huh-7 cells, the functional roles of the two forms of the viral protein, hepatitis delta antigen (HDAg), in HDV RNA replication are similar for both genotypes I and III; the short form is required for RNA replication, while the long form inhibits replication. For both genotypes, HDAg was able to support replication of RNAs of the same genotype that were mutated so as to be defective for HDAg production. Surprisingly, however, neither genotype I nor genotype III HDAg was able to support replication of such mutated RNAs of the other genotype. The inability of genotype III HDAg to support replication of genotype I RNA could have been due to a weak interaction between the RNA and HDAg. The clear genotype-specific activity of HDAg in supporting HDV RNA replication confirms the original categorization of HDV sequences in three genotypes and further suggests that these should be referred to as types (i.e., HDV-I and HDV-III) rather than genotypes.

scholarly journals Characterization of hepatitis delta antigen: specific binding to hepatitis delta virus RNA.

1990 ◽  
Vol 64 (9) ◽  
pp. 4051-4058 ◽  
Author(s):  
J H Lin ◽  
M F Chang ◽  
S C Baker ◽  
S Govindarajan ◽  
M M Lai
Keyword(s):  
Hepatitis Delta Virus ◽  
Specific Binding ◽  
Hepatitis Delta ◽  
Delta Antigen ◽  
Virus Rna ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

scholarly journals Hepatitis Delta Antigen Requires a Minimum Length of the Hepatitis Delta Virus Unbranched Rod RNA Structure for Binding

2009 ◽  
Vol 83 (9) ◽  
pp. 4548-4556 ◽  
Author(s):  
Dawn A. Defenbaugh ◽  
Matthew Johnson ◽  
Renxiang Chen ◽  
Ying Yi Zheng ◽  
John L. Casey
Keyword(s):  
Hepatitis Delta Virus ◽  
Rna Binding ◽  
Specific Binding ◽  
Micrococcal Nuclease ◽  
Minimum Length ◽  
Hepatitis Delta ◽  
Ribonucleoprotein Complexes ◽  
Delta Antigen ◽  
Hepatitis Delta Antigen ◽  
Delta Virus

ABSTRACT Hepatitis delta virus (HDV) is a subviral pathogen that increases the severity of liver disease caused by hepatitis B virus. Both the small circular RNA genome and its complement, the antigenome, form a characteristic unbranched rod structure in which approximately 70% of the nucleotides are base paired. These RNAs are associated with the sole virally encoded protein, hepatitis delta antigen (HDAg), in infected cells; however, the nature of the ribonucleoprotein complexes (RNPs) is not well understood. Previous analyses of binding in vitro using native, bacterially expressed HDAg have been hampered by a lack of specificity for HDV RNA. Here, we show that removal of the C-terminal 35 amino acids of HDAg yields a native, bacterially expressed protein, HDAg-160, that specifically binds HDV unbranched rod RNA with high affinity. In an electrophoretic mobility shift assay, this protein produced a discrete, micrococcal nuclease-resistant complex with an ∼400-nucleotide (nt) segment of HDV unbranched rod RNA. Binding occurred with several segments of HDV RNA, although with various affinities and efficiencies. Analysis of the effects of deleting segments of the unbranched rod indicated that binding did not require one or two specific binding sites within these RNA segments. Rather, a minimum-length HDV RNA unbranched rod approximately 311 nt was essential for RNP formation. The results are consistent with a model in which HDAg binds HDV unbranched rod RNA as multimers of fixed size rather than as individual subunits.

scholarly journals Transcription of Hepatitis Delta Antigen mRNA Continues throughout Hepatitis Delta Virus (HDV) Replication: a New Model of HDV RNA Transcription and Replication

1998 ◽  
Vol 72 (7) ◽  
pp. 5449-5456 ◽  
Author(s):  
Lucy E. Modahl ◽  
Michael M. C. Lai
Keyword(s):  
Hepatitis Delta Virus ◽  
Rna Synthesis ◽  
Rna Replication ◽  
Hepatitis Delta ◽  
New Model ◽  
Delta Antigen ◽  
Hepatitis Delta Antigen ◽  
Genomic Length ◽  
Polyadenylated Mrna ◽  
Delta Virus

ABSTRACT Hepatitis delta virus (HDV) replicates by RNA-dependent RNA synthesis according to a double rolling circle model. Also synthesized during replication is a 0.8-kb, polyadenylated mRNA encoding the hepatitis delta antigen (HDAg). It has been proposed that this mRNA species represents the initial product of HDV RNA replication; subsequent production of genomic-length HDV RNA relies on suppression of the HDV RNA polyadenylation signal by HDAg. However, this model was based on studies which required the use of an HDV cDNA copy to initiate HDV RNA replication in cell culture, thus introducing an artificial requirement for DNA-dependent RNA synthesis. We have now used an HDV cDNA-free RNA transfection system and a method that we developed to detect specifically the mRNA species transcribed from the HDV RNA template. We established that this polyadenylated mRNA is 0.8 kb in length and its 5′ end begins at nucleotide 1631. Surprisingly, kinetic studies showed that this mRNA continued to be synthesized even late in the viral replication cycle and that the mRNA and the genomic-length RNA increased in parallel, even in the presence of HDAg. Thus, a switch from production of the HDAg mRNA to the full-length HDV RNA does not occur in this system, and suppression of the polyadenylation site by HDAg may not significantly regulate the synthesis of the HDAg mRNA, as previously proposed. These findings reveal novel insights into the mechanism of HDV RNA replication. A new model of HDV RNA replication and transcription is proposed.

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