Base Pairing between the 5′ Half of ε and a
            <i>cis</i>
            -Acting Sequence, Φ, Makes a Contribution to the Synthesis of Minus-Strand DNA for Human Hepatitis B Virus

Abraham, Teresa M.; Loeb, Daniel D.

doi:10.1128/jvi.80.9.4380-4387.2006

Cited by 42 publications

(58 citation statements)

References 28 publications

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“…Most of is partially complementary to the 5´ half of ε. Mutations affecting the base-pairing potential between the two sequences can, indeed, influence the efficiency of (-)-DNA synthesis [147] . Exactly how an ε-φ interaction would aid in primer translocation is unclear because base-pairing would simultaneously affect the proper ε structure, implying some temporal regulation.…”

Section: ( + ) -S T R a N D D N A S Y N T H E S I S A N D Circularizamentioning

confidence: 99%

“…via elongation initiation factor 4G (eIF-4G; large oval). More specifically, ε might base-pair with φ [146,147] , as indicated by the grey arrows. In such an arrangement, a small movement, rather than a big jump, of TP with the bound DNA primer (dashed outline) would suffice for specific translocation to DR1*.…”

Section: Dna Primer Translocation and (-)-Dna Completionmentioning

confidence: 99%

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Hepatitis B virus replication

Beck

Nassal

2007

WJG

413

382

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Hepadnaviruses, including human hepatitis B virus (HBV), replicate through reverse transcription of an RNA intermediate, the pregenomic RNA (pgRNA). Despite this kinship to retroviruses, there are fundamental differences beyond the fact that hepadnavirions contain DNA instead of RNA. Most peculiar is the initiation of reverse transcription: it occurs by protein-priming, is strictly committed to using an RNA hairpin on the pgRNA, ε, as template, and depends on cellular chaperones; moreover, proper replication can apparently occur only in the specialized environment of intact nucleocapsids. This complexity has hampered an in-depth mechanistic understanding. The recent successful reconstitution in the test tube of active replication initiation complexes from purified components, for duck HBV (DHBV), now allows for the analysis of the biochemistry of hepadnaviral replication at the molecular level. Here we review the current state of knowledge at all steps of the hepadnaviral genome replication cycle, with emphasis on new insights that turned up by the use of such cellfree systems. At this time, they can, unfortunately, not be complemented by three-dimensional structural information on the involved components. However, at least for the ε RNA element such information is emerging, raising expectations that combining biophysics with biochemistry and genetics will soon provide a powerful integrated approach for solving the many outstanding questions. The ultimate, though most challenging goal, will be to visualize the hepadnaviral reverse transcriptase in the act of synthesizing DNA, which will also have strong implications for drug development.© 2007 The WJG Press. All rights reserved. Dieter Glebe, PhD, Series EditorPO Box 2345, Beijing 100023, China World J Gastroenterol 2007 January 7; 13(1): 48-64 www.wjgnet.com World Journal of Gastroenterology ISSN 1007-9327 wjg@wjgnet.com © 2007 OVERVIEW OVER THE HEPADNAVIRAL GENOME REPLICATION CYCLEReplication of the hepadnaviral genome can broadly be divided into three phases ( Figure 1): (1) Infectious virions contain in their inner icosahedral core the genome as a partially double-stranded, circular but not covalently closed DNA of about 3.2 kb in length (relaxed circular, or RC-DNA); (2) upon infection, the RC-DNA is converted, inside the host cell nucleus, into a plasmid-like covalently closed circular DNA (cccDNA); (3) from the cccDNA, several genomic and subgenomic RNAs are transcribed by cellular RNA polymerase Ⅱ; of these, the pregenomic RNA (pgRNA) is selectively packaged into progeny capsids and is reverse transcribed by the co-packaged P protein into new RC-DNA genomes. Matured RC-DNA containing-but not immature RNA containingnucleocapsids can be used for intracellular cccDNA amplification, or be enveloped and released from the cell as progeny virions. Below we discuss these genome conversions, with emphasis on the reverse transcription step, and particularly its unique initiation mechanism. RC-DNA TO cccDNA CONVERSIONPersistent viral infections require tha...

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Section: ( + ) -S T R a N D D N A S Y N T H E S I S A N D Circularizamentioning

confidence: 99%

Section: Dna Primer Translocation and (-)-Dna Completionmentioning

confidence: 99%

Hepatitis B virus replication

Beck

Nassal

2007

WJG

413

382

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“…6d). Recent studies have shown that mutation of the lower bulge of the e structure (1858-1863) prevents the pol-e interaction (Abraham & Loeb, 2006;Oropeza & McLachlan, 2006). We co-transfected eGFP-pol and core with a vector expressing e mutated in this fashion (ppreC-/C e mutant) and found that core localized in the nucleus (Fig.…”

Section: Cytoplasmic Core Localization Requires Pol and Ementioning

confidence: 99%

Expression of viral polymerase and phosphorylation of core protein determine core and capsid localization of the human hepatitis B virus

Deroubaix

Osseman

Cassany

et al. 2015

Journal of General Virology

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Biopsies from patients show that hepadnaviral core proteins and capsids -collectively called core -are found in the nucleus and cytoplasm of infected hepatocytes. In the majority of studies, cytoplasmic core localization is related to low viraemia while nuclear core localization is associated with high viral loads. In order to better understand the molecular interactions leading to core localization, we analysed transfected hepatoma cells using immune fluorescence microscopy. We observed that expression of core protein in the absence of other viral proteins led to nuclear localization of core protein and capsids, while expression of core in the context of the other viral proteins resulted in a predominantly cytoplasmic localization. Analysis of which viral partner was responsible for cytoplasmic retention indicated that the HBx, surface proteins and HBeAg had no impact but that the viral polymerase was the major determinant. Further analysis revealed that e, an RNA structure to which the viral polymerase binds, was essential for cytoplasmic retention. Furthermore, we showed that core protein phosphorylation at Ser 164 was essential for the cytoplasmic core localization phenotype, which is likely to explain differences observed between individual cells.

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“…1B) (48,53,62,64). The specificity of this template switch is determined by base pairing between nucleotides in ε and nucleotides flanking the acceptor site ( and ) (2,3). Subsequently, the minus-strand DNA is elongated to produce a full-length minus-strand DNA [(Ϫ)DNA elongation] (Fig.…”

mentioning

confidence: 99%

The Arginine Clusters of the Carboxy-Terminal Domain of the Core Protein of Hepatitis B Virus Make Pleiotropic Contributions to Genome Replication

Lewellyn

Loeb

2011

J Virol

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The carboxy-terminal domain (CTD) of the core protein of hepatitis B virus is not necessary for capsid assembly. However, the CTD does contribute to encapsidation of pregenomic RNA (pgRNA). The contribution of the CTD to DNA synthesis is less clear. This is the case because some mutations within the CTD increase the proportion of spliced RNA to pgRNA that are encapsidated and reverse transcribed. The CTD contains four clusters of consecutive arginine residues. The contributions of the individual arginine clusters to genome replication are unknown. We analyzed core protein variants in which the individual arginine clusters were substituted with either alanine or lysine residues. We developed assays to analyze these variants at specific steps throughout genome replication. We used a replication template that was not spliced in order to study the replication of only pgRNA. We found that alanine substitutions caused defects at both early and late steps in genome replication. Lysine substitutions also caused defects, but primarily during later steps. These findings demonstrate that the CTD contributes to DNA synthesis pleiotropically and that preserving the charge within the CTD is not sufficient to preserve function.Hepatitis B virus (HBV) replicates its genome via reverse transcription of a pregenomic RNA (pgRNA). This process ultimately produces a double-stranded DNA with a relaxedcircular conformation (rcDNA) (for a review, see reference 57). A distinguishing feature of HBV reverse transcription is that it occurs within the viral capsid. In the absence of HBV capsids, the HBV polymerase (P) is not able to produce rcDNA genomes from pgRNAs (36,58,61). Observations like these suggest that the capsid participates in reverse transcription.HBV capsids are icosahedra with primarily Tϭ4 symmetry (9,16,19). Each capsid contains 120 dimers of the core protein (16,69), and capsids can assemble in the absence of other viral components (59). A region of the core protein that is a candidate for a role in reverse transcription is the carboxy-terminal region of 34 amino acids, known as the carboxy-terminal domain (CTD). Capsids without the CTD assemble normally but do not support genome replication (8,47,70). The CTD can be attached to the capsid interior at the 5-or quasi-6-fold axes (71). Also, the CTD can bind to nucleic acids in vitro (28). Nearly 50% of the residues in the CTD are arginines, which give the CTD a net positive charge. Fourteen of the sixteen arginines are grouped into four clusters of three or four (see Fig. 2A for the amino acid sequence). This arrangement is conserved among mammalian hepadnaviruses ( Fig. 2A), suggesting it is important for function. Serines at positions 155, 162, and 170 are also conserved and can be phosphorylated in HBV (17,21,40). However, it is not fully understood which features of the CTD (arginine clusters, serine phosphoacceptor sites, or other) contribute to genome replication. The goal of the present study is to understand how each arginine cluster contributes to genome replicat...

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Base Pairing between the 5′ Half of ε and a cis -Acting Sequence, Φ, Makes a Contribution to the Synthesis of Minus-Strand DNA for Human Hepatitis B Virus

Cited by 42 publications

References 28 publications

Hepatitis B virus replication

Hepatitis B virus replication

Expression of viral polymerase and phosphorylation of core protein determine core and capsid localization of the human hepatitis B virus

The Arginine Clusters of the Carboxy-Terminal Domain of the Core Protein of Hepatitis B Virus Make Pleiotropic Contributions to Genome Replication

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