1054-1061.)A utophagy is important for removing long-lived proteins and damaged organelles in cells. During autophagy, double-membrane vesicles form to sequester part of the cytoplasm. These double-membrane vesicles, also known as autophagosomes, subsequently fuse with lysosomes to form autolysosomes for the degradation of their contents for recycling. 1 Many genes that are important for autophagy have been identified. Among them is microtubule-associated protein light chain 3 (LC3), whose covalent linkage to phosphatidylethanolamine by the ubiquitin-activating enzyme E1-like protein Atg7 is necessary for the formation of autophagosomes. 2 Hepatitis C virus (HCV) is a positive-stranded RNA virus with a genome size of 9.6 Kb. Infection by this virus can lead to liver cirrhosis and hepatocellular carcinoma. Based on their genetic relatedness, different HCV isolates have been grouped into six major genotypes and many more subtypes. The HCV genome codes for a polyprotein, which is proteolytically cleaved to generate the mature protein products. 3 Recently, a cell culture system for efficient HCV propagation using the JFH1 strain, which belongs to HCV genotype 2a, has been developed. [4][5][6][7] In this system, the HCV JFH1 RNA or its derivative was transfected into human hepatoma cells to direct the replication and release of infectious HCV particles, which could then initiate the next round of infection. In this report, we use this HCV RNA transfection/ infection system to study HCV-host interactions. Our results indicate that HCV induces the accumulation of autophagosomes by activating unfolded protein response (UPR). However, HCV does not enhance autophagic protein degradation. Importantly, this induction of autophagosomes enhanced HCV replication. The persistent induction of the UPR and the Abbreviations: BAF, bafilomycin A1; DMEM, Dulbecco's modified Eagle's medium; DTT, dithiothreitol; ER, endoplasmic reticulum; GFP, green fluorescence protein; HCV, hepatitis C virus; LC3, mRNA, messenger RNA; PERK, siRNA, small interfering RNA; Tg, thapsigargin; UPR, unfolded protein response. From the

Autophagy is a catabolic process by which cells remove long-lived proteins and damaged organelles for recycling. Viral infections may also induce autophagic response. Here we show that hepatitis B virus (HBV), a pathogen that chronically infects ≈350 million people globally, can enhance autophagic response in cell cultures, mouse liver, and during natural infection. This enhancement of the autophagic response is not coupled by an increase of autophagic protein degradation and is dependent on the viral X protein, which binds to and enhances the enzymatic activity of phosphatidylinositol 3-kinase class III, an enzyme critical for the initiation of autophagy. Further analysis indicates that autophagy enhances HBV DNA replication, with minimal involvement of late autophagic vacuoles in this process. Our studies thus demonstrate that a DNA virus can use autophagy to enhance its own replication and indicate the possibility of targeting the autophagic pathway for the treatment of HBV patients.autophagy | hepatitis B virus DNA replication | hepatitis B virus X protein | PI3KC3A utophagy is a catabolic process by which long-lived proteins and damaged organelles are sequestered in the cytoplasm and removed for recycling. It is important for maintaining cellular homeostasis. During autophagy, membrane crescents appear in the cytoplasm. These membranes will eventually form a double-membrane structure known as autophagosomes, which will mature by fusing with lysosomes to form autolysosomes. The contents of autophagosomes will subsequently be degraded by lysosomal enzymes. Autophagy has also been implicated in innate and adaptive immune responses to the infection of microbial pathogens (1, 2). A number of viruses have been shown to induce autophagy, either completely or partially, and often with either a destructive or beneficial result to themselves. For examples, several single-stranded RNA viruses such as poliovirus, coronavirus, dengue virus, and hepatitis C virus all seem to induce the accumulation of autophagic vacuoles and use these membrane vesicles to benefit their replication (3-6). In contrast, other viruses such as herpes simplex virus-1 (HSV-1), cytomegalovirus (CMV), and Kaposi's sarcoma herpes virus (KSHV) have evolved mechanisms to suppress autophagy and, in the case of HSV-1, for its own survival (2).Hepatitis B virus (HBV) belongs to the Hepadnavirus family. This virus has a 3.2-kb circular and partially double-stranded DNA genome that contains four genes named S, C, P, and X genes. The S gene codes for the surface antigens (i.e., envelope proteins), the C gene codes for the core protein and a related protein termed precore protein, the P gene codes for the viral DNA polymerase, and the X gene codes for a multifunctional regulatory protein. After its synthesis, the core protein packages its own mRNA, which is also known as the pregenomic RNA (pgRNA), to form the core particle. The pgRNA will be converted to the DNA genome in the core particle by the viral DNA polymerase, which is also a reverse transcriptase...

Background: Hepatitis C virus (HCV) induces autophagosomes in its host cells. Results: The HCV RNA replication complex colocalizes with autophagosomes, which are induced by HCV via a Class III PI3K-independent pathway. Conclusion: HCV induces autophagosomes and uses their membranes for its RNA replication. Significance: The perturbation of the autophagic pathway by HCV may have important consequences in HCV pathogenesis.

Hepatitis B virus (HBV) is a hepatotropic virus that can cause severe liver diseases, including liver cirrhosis and hepatocellular carcinoma. This virus chronically infects approximately 350 million people in the world, causing significant morbidity and mortality. HBV is a small DNA virus with a partially double-stranded and circular DNA genome that has a length of about 3.2 kb. After the infection of hepatocytes, this DNA is repaired to form a covalently closed circular DNA (cccDNA) molecule, which then directs the transcription of viral mRNAs. The mRNA of the viral core protein is larger than the genome length. This core protein mRNA, which is also termed the pregenomic RNA (pgRNA), is packaged by the core protein to form the viral core particle. It is subsequently converted to the partially double-stranded viral genome by the viral RNA polymerase, which is also packaged in the core particle. The core particle subsequently interacts with the viral envelope proteins for the formation of the mature virion, which is then released from infected cells (for a review, see reference 1).Recently, we demonstrated that HBV can induce autophagy in cell cultures, in the mouse liver, and during natural infection

The role of autophagy in carcinogenesis is controversial and apparently complex. By using mice with hepatocyte-specific knockout of Atg5, a gene essential for autophagy, we longitudinally studied the role of autophagy in hepatocarcinogenesis. We found that impairing autophagy in hepatocytes would induce oxidative stress and DNA damage, followed by the initiation of hepatocarcinogenesis, which could be suppressed by the antioxidant N-acetylcysteine. Interestingly, these mice developed only benign tumors with no hepatocellular carcinoma (HCC), even after the treatment with diethylnitrosamine, which induced HCC in wild-type mice. The inability of mice to develop HCC when autophagy was impaired was associated with the induction of multiple tumor suppressors including p53. Further analysis indicated that the induction of p53 was associated with the DNA-damage response. Tumorigenesis studies using an established liver tumor cell line confirmed a positive role of autophagy in tumorigenesis and a negative role of p53 in this process when autophagy was impaired. Our studies thus demonstrate that autophagy is required to maintain healthy mitochondria and to reduce oxidative stress and DNA damage to prevent the initiation of hepatocarcinogenesis. However, once hepatocarcinogenesis has been initiated, its presence is also required to suppress the expression of tumor suppressors to promote the development of HCC. Autophagy (i.e., macroautophagy) is important for cells to remove protein aggregates and damaged organelles. Its dysfunction can cause a variety of diseases including cancers. 1,2 However, its role in carcinogenesis is apparently complex, as it has been shown in different reports to positively or negatively regulate carcinogenesis. 3,4 Autophagy apparently can function as a tumor suppressor, as the gene encoding Beclin-1, a component of the phosphatidylinositol-3-kinase class III (PI3KC3) complex that is essential for the initiation of autophagy, is often monoallelically deleted or mutated in breast, ovarian and prostate cancers. 5 Frameshift mutations in Atg2B, Atg5, Atg9B and Atg12 autophagy genes are also often found in gastric and colorectal cancers with microsatellite instability. 6 The tumor suppressor role of autophagy is further supported by the studies using mouse models. It has been shown that the monoallelic deletion of the Beclin-1 gene in mice induced tumor lesions in various tissues, 7 Atg4C-knockout (KO) mice had increased susceptibility to carcinogens for the development of fibrosarcomas 8 and the systemic mosaic KO of Atg5 and the liver-specific KO of Atg7 in mice led to the development of benign liver adenomas. 9,10 Autophagy has also been shown to promote tumor growth. It has been shown that autophagy can enhance the survival of tumor cells in the hypoxic regions of solid tumors. 11 It has also been shown that in cells expressing oncogenic Ras, autophagy is required to promote tumorigenesis by maintaining oxidative metabolism or facilitating glycolysis. 12,13 Moreover, it has also been demonstrate...

Runx2 and androgen receptor (AR) are master transcription factors with pivotal roles in bone metabolism and prostate cancer (PCa). We dissected AR-mediated repression of Runx2 in dihydrotestosterone (DHT)-treated osteoblastic and PCa cells using reporter assays and endogenous Runx2 target genes. Repression required DHT, but not AR's transactivation function, and was associated with nuclear colocalization of the two proteins. Runx2 and AR coimmunoprecipitated and interacted directly in glutathione-S-transferase pull-down assays. Interaction was ionic in nature. Intact AR DNA-binding domain (DBD) was necessary and sufficient for both interaction with Runx2 and its repression. Runx2 sequences required for interaction were the C-terminal 132 amino acid residues together with the Runt DBD. Runx2 DNA binding was abrogated by endogenous AR in chromatin immunoprecipitation assays and by recombinant AR-DBD in gel shift assays. Furthermore, AR caused increased nuclear mobility of Runx2 as indicated by faster fluorescence recovery after photobleaching. Thus, AR binds Runx2 and abrogates its binding to DNA and possibly to other nuclear components. Clinical relevance of our results was suggested by an inverse correlation between expression of AR-responsive prostate-specific antigen and osteocalcin genes in PCa biopsies. Given the tumor suppressor properties of Runx2, its repression by AR may constitute a mechanism of hormone carcinogenesis. Attenuation of Runx2 by AR in osteoblasts may play a role in skeletal metabolism: the bone-sparing effect of androgens is attributable, in part, to keeping Runx2 activity in check and preventing high-turnover bone disease such as seen after castration and in transgenic mice overexpressing Runx2 in osteoblasts.

Autophagy is a catabolic process that is important for the removal of damaged organelles and long-lived proteins for the maintenance of cellular homeostasis. It can also serve as innate immunity to remove intracellular microbial pathogens. A growing list of viruses has been shown to affect this cellular pathway. Some viruses suppress this pathway for their survival, while others enhance or exploit this pathway to benefit their replication. The effect of viruses on autophagy may also sensitize cells to death or enhance cell survival and play a critical role in viral pathogenesis. In this article, we review the relationships between different viruses and autophagy and discuss how these relationships may affect viruses and their host cells.

Autophagy is a catabolic process by which cells remove unwanted proteins and damaged organelles. It is important for maintaining cellular homeostasis and can also be used by cells to remove intracellular microbial pathogens. As such, some viruses such as herpes simplex virus-1 (HSV-1) have evolved mechanisms to suppress autophagy for their survival. In contrast, other viruses such as poliovirus, hepatitis C virus (HCV) and dengue viruses have instead evolved mechanisms to use this pathway to enhance their replication. Recently, we demonstrated that hepatitis B virus (HBV), a DNA virus that infects hepatocytes, could enhance and use autophagy for its DNA replication. This enhancement of autophagy is mediated by its X protein, which binds to and activates phosphatidylinositol-3-kinase class 3 (PI3KC3), an enzyme important for the initiation of autophagy. The persistent activation of autophagy in hepatocytes by HBV during chronic infection may play an important role in HBV pathogenesis.