The hepatitis B virus (HBV) is a major human pathogen, belonging to the family of hepadnaviruses, a group of small enveloped viruses with major liver tropism (47). The HBV genome consists of a relaxed, circular, partially double-stranded 3.2-kb DNA molecule. One of the striking features of HBV is that its replication involves reverse transcription of a greater-than-genome-length pregenomic RNA (3.5 kb) (22, 29). This reverse transcription process occurs exclusively in the core particle, which is assembled through complex interactions between pregenomic RNA, core protein, polymerase, and several cellular proteins (22, 29). The core particles containing the replicative intermediates are then transported back to the nucleus, for the establishment of a pool of covalently closed circular DNA, or to the endoplasmic reticulum to be released after association with the viral envelope, as infectious mature virions (for a review, see reference 22). HBV infection may lead to acute liver disease, chronic active hepatitis, liver cirrhosis, and hepatocellular carcinoma. Over 300 million people worldwide are estimated to be infected chronically by HBV and are therefore at risk of liver failure, cirrhosis, or hepatocellular carcinoma. The principal treatment for chronic hepatitis B involves the use of alpha interferon (IFN-α) or nucleoside analogs (9, 42). IFN-α belongs to the IFN-α/β system, which mediates antiviral, antiproliferative, immune, and other cellular effects (8). In humans, IFN-α antiviral action is mediated by the induction of at least three major proteins, 2′,5′-oligoadenylate synthetase, protein kinase R, and MxA. IFN-α likely acts by combining stimulation of the immune response and a direct viral effect. However, the specific mechanisms responsible for an improvement in HBV-related hepatitis following IFN treatment are not clearly understood. To date, IFN-α antiviral mechanisms against HBV have mainly been examined in vitro using hepatoma cell lines. These experiments showed that IFN-α brought about changes to the expression of viral antigens and/or steady-state levels of viral RNAs or replicative intermediates, depending on the experimental model employed (2, 4, 7, 17, 21, 26, 35, 36, 51, 54). Although the use of IFN-α has improved the treatment of chronically infected HBV patients, an effective reduction in virus load is only observed in 30% of treated patients. The molecular basis for resistance to IFN-α therapy is not clearly defined. However, studies have suggested that HBV may play a direct role in the development of resistance to endogenous or exogenous IFN. In vitro, HBV genome expression has been shown to reduce sensitivity to IFN, as measured by inhibition of the cytopathic effect of Sindbis virus challenge (30). HBV capsid and polymerase terminal proteins have been shown to reduce expression of the IFN-β and IFN-induced 6-16 genes, respectively (11, 52, 53). Furthermore, several in vivo studies have demonstrated a lack of IFN system activation in patients with acute or chronic hepatitis B. In particular, impaired induction of the IFN-inducible MxA protein was evidenced in acute and chronic HBV infection (10, 20). MxA is a 76-kDa GTPase protein belonging to the superfamily of large GTPases, which accumulate in the cytoplasm in response to IFN-α/β (16). In vitro, in different cellular models, or in vivo, in MxA transgenic mice, MxA protein is able to inhibit a broad spectrum of negative-stranded RNA viruses, including influenza virus, Thogoto virus, vesicular stomatitis virus, measles virus, and bunyavirus (12, 13, 33, 46, 57). Recently, antiviral activity has been demonstrated against a positive-stranded RNA virus, Semliki Forest virus (28). The mechanisms through which MxA is able to inhibit such a variety of viruses are yet to be precisely defined. Several studies have shown that MxA may act at different levels of the virus replication cycle, depending on the virus species and the cellular models used. Indeed, MxA is capable of blocking viral replication at primary transcriptional steps (Semliki Forest and vesicular stomatitis viruses) (28, 46) or following primary transcription (influenza virus) (32). In the case of measles virus, MxA seems to have an inhibitory effect on either viral RNA or glycoprotein synthesis, depending on the cellular model (43, 44). In a previous study, we showed evidence for HBV defective particles, characterized by a singly spliced HBV RNA, which had been encapsidated and retrotranscribed, giving rise to a defective HBV genome (49). We also demonstrated an association between those defective particles and the establishment of a chronic carrier state (37). In vitro, we showed that expression of this defective genome led to a reduction in the antiviral activity of IFN, as determined using the virus yield reduction assay, and that this modulation involved a selective inhibition of MxA protein induction via overexpression of the HBV capsid protein (38). This led us to suggest that MxA might play a major role in antiviral activity against HBV. The aim of the present study was to determine whether the antiviral spectrum of the MxA protein extended to cover HBV. We therefore established HuH7 cell lines stably expressing the MxA protein and performed transient-transfection experiments using an HBV-expressing plasmid. We found that MxA inhibited HBV replication through a significant reduction in the synthesis of viral proteins, cytoplasmic RNAs, and DNA replicative intermediates. We demonstrate that MxA antiviral action against HBV occurs, partly at least, at a posttranscriptional level, by inhibiting the nuclear export of viral RNAs.