Yi Hsuan Hsieh

Hepatitis B virus Pre-S2 mutant surface antigen induces degradation of cyclin-dependent kinase inhibitor p27Kip1 through c-jun activation domain-binding protein 1

The hepatitis B virus (HBV) large surface antigen (LHBS) mutant with deletion at the pre-S2 region accumulates in endoplasmic reticulum (ER) and is associated with HBV-induced hepatocellular carcinogenesis. In this study, we found that the pre-S2 LHBS mutant directly interacts with the Jun activation domain–binding protein 1 (JAB1). Association of pre-S2 LHBS with JAB1 dissociated JAB1 from the JAB1/IRE1 complex in ER. The free (active) JAB1 then translocated into cell nuclei and rendered the Cdk inhibitor p27Kip1 to cytosolic proteasome for degradation. The pre-S2 LHBS mutant induced hyperphosphorylation of tumor suppressor retinoblastoma (RB) via cyclin-dependent kinase 2 (Cdk2), a downstream molecule regulated by p27Kip1. This effect is independent of the ER stress signaling pathway. The transgenic mice carrying the pre-S2 mutant LHBS gene also exhibited Cdk2 activation, p27Kip1 degradation, as well as RB hyperphosphorylation. The mouse hepatocytes exhibited morphologic abnormalities such as chromatin condensation, multinucleation, and dysplasia of hepatocytes. In summary, the pre-S2 LHBS mutant causes p27Kip1 degradation through direct interaction with JAB1. The pre-S2 mutant LHBS is suggested to be a potential oncoprotein for HBV-related hepatocellular carcinoma. (Mol Cancer Res 2007;5(10):1063–72)

Genomic instability caused by hepatitis B virus: into the hepatoma inferno.

Chronic hepatitis B virus (HBV) infection is an important cause of hepatocellular carcinoma (HCC) worldwide, especially in Asia. HBV induces HCC through multiple oncogenic pathways. Hepatitis-induced hepatocyte inflammation and regeneration stimulates cell proliferation. The interplay between the viral and host factors activates oncogenic signaling pathways and triggers cell transformation. In this review, we summarize previous studies, which reported that HBV induces host genomic instability and that HBV-induced genomic instability is a significant factor that accelerates carcinogenesis. The various types of genomic changes in HBV-induced HCC--chromosomal instability, telomere attrition, and gene-level mutations--are reviewed. In addition, the two viral factors, HBx and the pre-S2 mutant large surface antigen, are discussed for their roles in promoting genomic instability as their main features as viral oncoproteins.

Hepatitis B virus pre-S2 mutant large surface protein inhibits DNA double-strand break repair and leads to genome instability in hepatocarcinogenesis

Although hepatitis B virus (HBV) has been established to cause hepatocellular carcinoma (HCC), the exact mechanism remains to be clarified. Type II ground glass hepatocytes (GGHs) harbouring the HBV pre‐S2 mutant large surface protein (LHBS) have been recognized as a morphologically distinct hallmark of HCC in the advanced stages of chronic HBV infection. Considering its preneoplastic nature, we hypothesized that type II GGH may exhibit high genomic instability, which is important for the carcinogenic process in chronic HBV carriers. In this study we found that pre‐S2 mutant LHBS directly interacted with importin α1, the key factor that recognizes cargos undergoing nuclear transportation mediated by the importin α/β‐associated nuclear pore complex (NPC). By interacting with importin α1, which inhibits its function as an NPC factor, pre‐S2 mutant LHBS blocked nuclear transport of an essential DNA repair and recombination factor, Nijmegen breakage syndrome 1 (NBS1), upon DNA damage, thereby delaying the formation of nuclear foci at the sites of DNA double‐strand breaks (DSBs). Pre‐S2 mutant LHBS was also found to block NBS1‐mediated homologous recombination repair and induce multi‐nucleation of cells. In addition, pre‐S2 mutant LHBS transgenic mice showed genomic instability, indicated by increased global gene copy number variations (CNVs), which were significantly higher than those in hepatitis B virus X mice, indicating that pre‐S2 mutant LHBS is the major viral oncoprotein inducing genomic instability in HBV‐infected hepatocytes. Consistently, the human type II GGHs in HCC patients exhibited increased DNA DSBs representing significant genomic instability. In conclusion, type II GGHs harbouring HBV pre‐S2 mutant oncoprotein represent a high‐risk marker for the loss of genome integrity in chronic HBV carriers and explain the complex chromosome changes in HCCs. Mouse array CGH raw data: GEO Accession No. GSE61378 (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE61378) Copyright

Histone deacetylase inhibitor suberoylanilide hydroxamic acid suppresses the pro-oncogenic effects induced by hepatitis b virus pre-S2 mutant oncoprotein and represents a potential chemopreventive agent in high-risk chronic HBV patients

Chronic hepatitis B virus (HBV) infection is the major cause of hepatocellular carcinoma (HCC). The pre-S(2) mutant large HBV surface antigen (LHBS) in type II ground glass hepatocytes (GGHs) has been recognized as an emerging viral oncoprotein; it directly interacts with the c-Jun activation domain-binding protein 1 (JAB1) and subsequently causes hyperphosphorylation of the tumor-suppressor retinoblastoma and, consequently, leads to disturbed cell cycle progression. The interaction of the pre-S(2) mutant LHBS with JAB1 could provide a potential target for chemoprevention. In this study, we found that the preneoplastic type II GGHs showed a significant decrease of the cyclin-dependent kinase inhibitor p27(Kip1), which serves as a marker for pre-S(2) mutant-JAB1 complex formation. The histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) elevated expression of the tumor-suppressor thioredoxin-binding protein 2 (TBP2), which subsequently enhanced the JAB1-TBP2 interaction and abolished the pre-S(2) mutant LHBS-induced degradation of p27(Kip1), which, in turn, recovered the normal cell cycle checkpoint. The pre-S(2) mutant LHBS-induced pro-oncogenic effects: increased cell proliferation, nuclear/cytoplasmic ratio and proliferating cell nuclear antigen expression, were all greatly ameliorated after SAHA treatments, which suggested SAHA as a promising chemopreventive agent for the pre-S(2) mutant oncoprotein-induced HCC. In conclusion, this study provides the mechanism of histone deacetylase (HDAC) inhibitor in preventing the pre-S(2) mutant-induced oncogenic phenotype. The HDAC inhibitor SAHA is therefore a potential chemopreventive agent for high-risk chronic HBV patients who may develop HCC.

A pre-S gene chip to detect pre-S deletions in hepatitis B virus large surface antigen as a predictive marker for hepatoma risk in chronic hepatitis B virus carriers

BackgroundChronic hepatitis B virus (HBV) infection is an important cause of hepatocellular carcinoma (HCC) worldwide. The pre-S1 and -S2 mutant large HBV surface antigen (LHBS), in which the pre-S1 and -S2 regions of the LHBS gene are partially deleted, are highly associated with HBV-related HCC.MethodsThe pre-S region of the LHBS gene in two hundred and one HBV-positive serum samples was PCR-amplified and sequenced. A pre-S oligonucleotide gene chip was developed to efficiently detect pre-S deletions in chronic HBV carriers. Twenty serum samples from chronic HBV carriers were analyzed using the chip.ResultsThe pre-S deletion rates were relatively low (7%) in the sera of patients with acute HBV infection. They gradually increased in periods of persistent HBV infection: pre-S mutation rates were 37% in chronic HBV carriers, and as high as 60% in HCC patients. The Pre-S Gene Chip offers a highly sensitive and specific method for pre-S deletion detection and is less expensive and more efficient (turnaround time 3 days) than DNA sequencing analysis.ConclusionThe pre-S1/2 mutants may emerge during the long-term persistence of the HBV genome in carriers and facilitate HCC development. Combined detection of pre-S mutations, other markers of HBV replication, and viral titers, offers a reliable predictive method for HCC risks in chronic HBV carriers.

Essential role of β-human 8-oxoguanine DNA glycosylase 1 in mitochondrial oxidative DNA repair.

8‐Oxoguanine (8‐OG) is the major mutagenic base lesion in DNA caused by reactive oxygen species (ROS) and accumulates in both nuclear and mitochondrial DNA (mtDNA). In humans, 8‐OG is primarily removed by human 8‐OG DNA glycosylase 1 (hOGG1) through the base excision repair (BER) pathway. There are two major hOGG1 isoforms, designated α‐ and β‐hOGG1, generated by alternative splicing, and they have distinct subcellular localization: cell nuclei and mitochondria, respectively. Using yeast two‐hybrid screening assays, we found that β‐ but not α‐hOGG1 directly interacts with the mitochondrial protein NADH:ubiquinone oxidoreductase 1 beta subcomplex 10 (NDUFB10), an integral factor in Complex 1 on the mitochondrial inner membrane. Using coimmunoprecipitation and immunofluorescence studies, we found that this interaction was greatly increased by hydrogen peroxide‐induced oxidative stress, suggesting that β‐ but not α‐hOGG1 is localized in the mitochondrial inner membrane. Analyses of nuclear and mtDNA damage showed that the β‐ but not α‐ hogg1 knockdown (KD) cells were severely defective in mitochondrial BER, indicating an essential requirement of β‐hOGG1 for mtDNA repair. β‐hogg1 KD cells were also found to be mildly deficient in Complex I activity, suggesting that β‐hOGG1 is an accessory factor for the mitochondrial integral function for ATP synthesis. In summary, our findings define β‐hOGG1 as an important factor for mitochondrial BER and as an accessory factor in the mitochondrial Complex I function. Mol. Mutagen. 2013.

Streptococcal Pyrogenic Exotoxin B Cleaves Human S-Adenosylhomocysteine Hydrolase and Induces Hypermethioninemia

Group A Streptococcus is a common pathogen that causes pharyngitis, impetigo, myositis, and lethal streptococcal toxic shock syndrome. Streptococcal pyrogenic exotoxin B (SPE B) is strongly associated with the severity of disease. SPE B is a cysteine protease and matures itself by autocatalysis. We found that SPE B was directly associated with human S-adenosylhomocysteine hydrolase (AdoHcyase), an essential factor for a delayed-type immune response. AdoHcyase protein levels and enzymatic activities were significantly higher in human cells infected with the Streptococcus pyogenes SW510 speB mutant strain than in cells infected with the NZ131 wild-type strain. SPE B also inactivated AdoHcyase, shown by a decrease in homocysteine, the main product of AdoHcyase. We found that in vivo and in vitro, SPE B induced hypermethioninemia, which is caused by an AdoHcyase defect. We also found that AdoHcyase is a substrate of SPE B cysteine protease. SPE B, therefore, potentially causes immunosuppression by cleaving AdoHcyase.