Because the replication of hepatitis B virus (HBV) proceeds via an obligatory reverse transcription step in the viral capsid, cDNA is potentially vulnerable to editing by cytidine deaminases of the APOBEC3 family. To date only two edited HBV genomes, referred to as G 3 A hypermutants, have been described in vivo. Recent work suggested that HBV replication was indeed restricted by APOBEC3G but by a mechanism other than editing. The issue of restriction has been explored by using a sensitive PCR method allowing differential amplification of AT-rich DNA. G 3 A hypermutated HBV genomes were recovered from transfection experiments involving APOBEC3B, -3C, -3F, and -3G indicating that all four enzymes were able to extensively deaminate cytidine residues in minus-strand DNA. Unexpectedly, three of the four enzymes (APOBEC3B, -3F, and -3G) deaminated HBV plus-strand DNA as well. From the serum of two of four patients with high viremia, G 3 A hypermutated genomes were recovered at a frequency of Ϸ10 ؊4 , indicating that they are, albeit relatively rare, part of the natural cycle of HBV infection. These findings suggest that human APOBEC3 enzymes can impact HBV replication via cytidine deamination.hypermutation ͉ DNA editing G 3 A hypermutated retroviral genomes result from the editing of nascent DNA by APOBEC3 cytidine deaminases (1-7). This was originally demonstrated for HIV-1 and the APOBEC3G member of a cluster of seven genes (3A-3H) on human chromosome 22 (8, 9). Editing occurs on the background of a ⌬vif genome (1-5, 10). The HIV Vif protein prevents packaging of either APOBEC3F or -3G resulting in their ubiquitination and degradation by the proteasomal pathway (11)(12)(13)(14). Editing of cytidine residues in neo-synthesized minusstrand cDNA results in the formation of multiple uracil residues that are read as T during plus-strand synthesis. Albeit referred to as G 3 A hypermutants by reference to the viral plus strand, mechanistically the action is occurring on the minus strand and independently of reverse transcriptase (7). Up to 60% of G residues in a lentiviral genome can be substituted resulting in the total loss of information (15-17). As such, APOBEC3F and -3G constitute a powerful restriction mechanism to reverse transcription. Retroviruses have either to avoid replication in cells expressing APOBEC3 molecules or else evolve a mechanism that neutralizes their effect. The vif gene of human and primate lentiviruses, as well as their homologues in most of the other lentiviruses, reflect the latter solution.Hepatitis B viruses (HBVs) replicate via an obligate reverse transcription step occurring in a capsid structure close to the endoplasmic reticulum. A pair of G 3 A hypermutated genomes were identified in the blood of a chronically infected patient yet have remained unique despite a burgeoning database (18). Recent reports demonstrated that HBV replication could be strongly restricted by APOBEC3G and -3F in an experimental setting (20,21). Restriction was highlighted by a strong reduction in the proportion o...

Hypermutations in hepatitis B virus (HBV) DNA by APOBEC3 cytidine deaminases have been detected in vitro and in vivo, and APOBEC3G (A3G) and APOBEC3F (A3F) have been shown to inhibit the replication of HBV in vitro, but the presumably low or even absent hepatic expression of these enzymes has raised the question as to their physiological impact on HBV replication. We show that normal human liver expresses the mRNAs of APOBEC3B (A3B), APOBEC3C (A3C), A3F, and A3G. In primary human hepatocytes, interferon alpha (IFN-␣) stimulated the expression of these cytidine deaminases up to 14-fold, and the mRNAs of A3G, A3F, and A3B reached expression levels of 10%, 3%, and 3%, respectively, relative to GAPDH mRNA abundance. On transfection, the full-length protein A3B L inhibited HBV replication in vitro as efficiently as A3G or A3F, whereas the truncated splice variant A3B S and A3C had no effect. A3B L and A3B S were detected predominantly in the nucleus of uninfected cells; however, in HBV-expressing cells both proteins were found also in the cytoplasm and were associated with HBV viral particles, similarly to A3G and A3F. T he hepatitis B virus (HBV) infects more than 350 million people worldwide and is a leading cause of end-stage liver disease and of hepatocellular carcinoma. 1 HBV is non-cytopathic for hepatocytes; however, most newly HBV infected adult patients develop acute hepatitis because of a strong immune response that clears HBV from the liver, whereas approximately 5% of newly HBV-infected adult patients generate insufficient immunity and become chronically infected. 1,2 Administration of interferon alpha (IFN-␣) is a mainstay of therapy for chronically HBV-infected patients. 2 Interferons restrict the replication of HBV by inducing the expression of antiviral proteins that inhibit the formation of replication-competent HBV nucleocapsids, and ultimately can result in the resolution of the chronic HBV infection. 2-7 HBV and other hepadnaviruses replicate their partially double-stranded DNA genome within cytoplasmic core particles by reverse transcription of encapsidated pregenomic RNA and thus are related to retroviruses. 8,9 The cytidine deaminase APOBEC3G (A3G), which is encoded within a cluster of seven related editing enzymes (APOBEC3A-G) on chromosome 22, provides broad innate immunity against exogenous and endogenous retroelements. 10-16 Encapsidated into the retroviral particle A3G deaminates dCs of the retroviral minus strand

In the absence of the viral vif gene, human immunodeficiency virus (HIV) may be restricted by the APOBEC3G gene on chromosome 22. The role of the HIV Vif protein is to exclude host cell APOBEC3G from the budding virion. As APOBEC3G shows sequence homology to cytidine deaminases, it is presumed that in the absence of Vif, cytidine residues in the cDNA are deaminated yielding uracil. It is not known if additional proteins mediate APOBEC3G function or if deamination occurs in concert with reverse transcription. This report describes an in vitro assay showing that Baculovirus derived APOBEC3G alone extensively deaminates cDNA independently of reverse transcriptase. It reproduces the dinucleotide context typical of G --> A hypermutants derived from a Delta(vif) virus. By using an RNaseH- form of reverse transcriptase, it was shown that the cDNA has to be free of its RNA template to allow deamination. APOBEC3G deamination of dC or dCTP was not detected. In short, APOBEC3G is a single-stranded DNA cytidine deaminase capable of restricting retroviral replication.

Our study reveals new knowledge on the mechanisms underlying the succession of yeasts during wine fermentations. This knowledge may be of importance when creating defined, mixed starter cultures for the controlled production of wines with a wide range of flavour compositions.