Investigation of biochemical, enzymatic, and metagenomic profiles of garbage enzymes and its application in dumping site leachate treatment.

The current article focuses on the preparation and characterization of garbage enzyme (GE) and explores its applications in treating leachate. GE is prepared from fruit and vegetable wastes and characterized via analysis of metabolites, carbohydrates, proteins, antioxidants, and enzymatic activities. This study extends our understanding of GE by reporting the presence of various metabolites. Moreover, a metagenomic analysis of GE is presented, shedding light on the microbial diversity. Firmicutes emerged as the dominant phylum, surpassing other phyla, including Proteobacteria and Actinobacteria. When exploring the potential for leachate treatment, the results indicate that vegetable GE shows 68% reduction in COD (chemical oxygen demand) and 39% reduction in ammoniacal nitrogen. Similarly, non-citrus GE also showed 64% reduction in COD and a 37% reduction in ammoniacal nitrogen, followed by citrus GE with a 33% reduction in COD and a 34% reduction in ammoniacal nitrogen compared to the control.

Virus-associated neuroendocrine cancers: Pathogenesis and current therapeutics.

Neuroendocrine neoplasms (NENs) comprise malignancies involving neuroendocrine cells that often lead to fatal pathological conditions. Despite escalating global incidences, NENs still have poor prognoses. Interestingly, research indicates an intricate association of tumor viruses with NENs. However, there is a dearth of comprehension of the complete scenario of NEN pathophysiology and its precise connections with the tumor viruses. Interestingly, several cutting-edge experiments became helpful for further screening of NET for the presence of polyomavirus, Human papillomavirus (HPV), Kaposi sarcoma-associated herpesvirus (KSHV), Epstein Barr virus (EBV), etc. Current research on the neuroendocrine tumor (NET) pathogenesis provides new information concerning their molecular mechanisms and therapeutic interventions. Of note, scientists observed that metastatic neuroendocrine tumors still have a poor prognosis with a palliative situation. Different oncolytic vector has already demonstrated excellent efficacies in clinical studies. Therefore, oncolytic virotherapy or virus-based immunotherapy could be an emerging and novel therapeutic intervention. In-depth understanding of all such various aspects will aid in managing, developing early detection assays, and establishing targeted therapeutic interventions for NENs concerning tumor viruses. Hence, this review takes a novel approach to discuss the dual role of tumor viruses in association with NENs' pathophysiology as well as its potential therapeutic interventions.

Comprehensive role of SARS-CoV-2 spike glycoprotein in regulating host signaling pathway.

Since the outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, global public health and the economy have suffered unprecedented damage. Based on the increasing related literature, the characteristics and pathogenic mechanisms of the virus, and epidemiological and clinical features of the disease are being rapidly discovered. The spike glycoprotein (S protein), as a key antigen of SARS-CoV-2 for developing vaccines, antibodies, and drug targets, has been shown to play an important role in viral entry, tissue tropism, and pathogenesis. In this review, we summarize the molecular mechanisms of interaction between S protein and host factors, especially receptor-mediated viral modulation of host signaling pathways, and highlight the progression of potential therapeutic targets, prophylactic and therapeutic agents for prevention and treatment of SARS-CoV-2 infection.

STAT3 in Tumor-Associated Myeloid Cells: Multitasking to Disrupt Immunity.

Myeloid immune cells, such as dendritic cells, monocytes, and macrophages, play a central role in the generation of immune responses and thus are often either disabled or even hijacked by tumors. These new tolerogenic activities of tumor-associated myeloid cells are controlled by an oncogenic transcription factor, signal transducer and activator of transcription 3 (STAT3). STAT3 multitasks to ensure tumors escape immune detection by impairing antigen presentation and reducing production of immunostimulatory molecules while augmenting the release of tolerogenic mediators, thereby reducing innate and adaptive antitumor immunity. Tumor-associated myeloid cells and STAT3 signaling in this compartment are now commonly recognized as an attractive cellular target for improving efficacy of standard therapies and immunotherapies. Hereby, we review the importance and functional complexity of STAT3 signaling in this immune cell compartment as well as potential strategies for cancer therapy.

The regulatory role of protein phosphorylation in human gammaherpesvirus associated cancers.

Activation of specific sets of protein kinases by intracellular signal molecules has become more and more apparent in the past decade. Phosphorylation, one of key posttranslational modification events, is activated by kinase or regulatory protein and is vital for controlling many physiological functions of eukaryotic cells such as cell proliferation, differentiation, malignant transformation, and signal transduction mediated by external stimuli. Moreovers, the reversible modification of phosphorylation and dephosphorylation can result in different features of the target substrate molecules including DNA binding, protein-protein interaction, subcellular location and enzymatic activity, and is often hijacked by viral infection. Epstein-Barr virus (EBV) and Kaposi's sarcomaassociated herpesvirus (KSHV), two human oncogenic gamma-herpesviruses, are shown to tightly associate with many malignancies. In this review, we summarize the recent progresses on understanding of molecular properties and regulatory modes of cellular and viral proteins phosphorylation influenced by these two tumor viruses, and highlight the potential therapeutic targets and strategies against their related cancers.

An essential EBV latent antigen 3C binds Bcl6 for targeted degradation and cell proliferation.

The latent EBV nuclear antigen 3C (EBNA3C) is required for transformation of primary human B lymphocytes. Most mature B-cell malignancies originate from malignant transformation of germinal center (GC) B-cells. The GC reaction appears to have a role in malignant transformation, in which a major player of the GC reaction is Bcl6, a key regulator of this process. We now demonstrate that EBNA3C contributes to B-cell transformation by targeted degradation of Bcl6. We show that EBNA3C can physically associate with Bcl6. Notably, EBNA3C expression leads to reduced Bcl6 protein levels in a ubiquitin-proteasome dependent manner. Further, EBNA3C inhibits the transcriptional activity of the Bcl6 promoter through interaction with the cellular protein IRF4. Bcl6 degradation induced by EBNA3C rescued the functions of the Bcl6-targeted downstream regulatory proteins Bcl2 and CCND1, which resulted in increased proliferation and G1-S transition. These data provide new insights into the function of EBNA3C in B-cell transformation during GC reaction, and raises the possibility of developing new targeted therapies against EBV-associated cancers.

WITHDRAWN: Curcumin shifts RAS-induced pro-proliferative MEK/ERK-signaling toward pro-apoptotic p38MAPK/JNK1-signaling, triggering p53 activation and apoptosis.

This article has been withdrawn by the authors. A mistake was made during the preparation of Fig 1C, NKE panel. The Western blot data shown for p-ERK1/2 and actin are not from this set, but rather a similar set of data from a different experiment. The authors apologize to the readers.

Crocetin exploits p53-induced death domain (PIDD) and FAS-associated death domain (FADD) proteins to induce apoptosis in colorectal cancer.

Tumor suppressor p53 preserves the genomic integrity by restricting anomaly at the gene level. The hotspots for mutation in half of all colon cancers reside in p53. Hence, in a p53-mutated cellular milieu targeting cancer cells may be achievable by targeting the paralogue(s) of p53. Here we have shown the effectiveness of crocetin, a dietary component, in inducing apoptosis of colon cancer cells with varying p53 status. In wild-type p53-expressing cancer cells, p53 in one hand transactivates BAX and in parallel up-regulates p53-induced death domain protein (PIDD) that in turn cleaves and activates BID through caspase-2. Both BAX and t-BID converge at mitochondria to alter the transmembrane potential thereby leading to caspase-9 and caspase-3-mediated apoptosis. In contrast, in functional p53-impaired cells, this phytochemical exploits p53-paralogue p73, which up-regulates FAS to cleave BID through FAS-FADD-caspase-8-pathway. These findings not only underline the phenomenon of functional switch-over from p53 to p73 in p53-impaired condition, but also validate p73 as a promising and potential target for cancer therapy in absence of functional p53.

EBV Nuclear Antigen 3C Mediates Regulation of E2F6 to Inhibit E2F1 Transcription and Promote Cell Proliferation.

Epstein-Barr virus (EBV) is considered a ubiquitous herpesvirus with the ability to cause latent infection in humans worldwide. EBV-association is evidently linked to different types of human malignancies, mainly of epithelial and lymphoid origin. Of interest is the EBV nuclear antigen 3C (EBNA3C) which is critical for EBV-mediated immortalization. Recently, EBNA3C was shown to bind the E2F1 transcription regulator. The E2F transcription factors have crucial roles in various cellular functions, including cell cycle, DNA replication, DNA repair, cell mitosis, and cell fate. Specifically, E2F6, one of the unique E2F family members, is known to be a pRb-independent transcription repressor of E2F-target genes. In our current study, we explore the role of EBNA3C in regulating E2F6 activities. We observed that EBNA3C plays an important role in inducing E2F6 expression in LCLs. Our study also shows that EBNA3C physically interacts with E2F6 at its amino and carboxy terminal domains and they form a protein complex in human cells. In addition, EBNA3C stabilizes the E2F6 protein and is co-localized in the nucleus. We also demonstrated that both EBNA3C and E2F6 contribute to reduction in E2F1 transcriptional activity. Moreover, E2F1 forms a protein complex with EBNA3C and E2F6, and EBNA3C competes with E2F1 for E2F6 binding. E2F6 is also recruited by EBNA3C to the E2F1 promoter, which is critical for EBNA3C-mediated cell proliferation. These results demonstrate a critical role for E2F family members in EBV-induced malignancies, and provide new insights for targeting E2F transcription factors in EBV-associated cancers as potential therapeutic intervention strategies.

The Modulation of Apoptotic Pathways by Gammaherpesviruses.

Apoptosis or programmed cell death is a tightly regulated process fundamental for cellular development and elimination of damaged or infected cells during the maintenance of cellular homeostasis. It is also an important cellular defense mechanism against viral invasion. In many instances, abnormal regulation of apoptosis has been associated with a number of diseases, including cancer development. Following infection of host cells, persistent and oncogenic viruses such as the members of the Gammaherpesvirus family employ a number of different mechanisms to avoid the host cell's "burglar" alarm and to alter the extrinsic and intrinsic apoptotic pathways by either deregulating the expressions of cellular signaling genes or by encoding the viral homologs of cellular genes. In this review, we summarize the recent findings on how gammaherpesviruses inhibit cellular apoptosis via virus-encoded proteins by mediating modification of numerous signal transduction pathways. We also list the key viral anti-apoptotic proteins that could be exploited as effective targets for novel antiviral therapies in order to stimulate apoptosis in different types of cancer cells.

The Role of Gammaherpesviruses in Cancer Pathogenesis.

Worldwide, one fifth of cancers in the population are associated with viral infections. Among them, gammaherpesvirus, specifically HHV4 (EBV) and HHV8 (KSHV), are two oncogenic viral agents associated with a large number of human malignancies. In this review, we summarize the current understanding of the molecular mechanisms related to EBV and KSHV infection and their ability to induce cellular transformation. We describe their strategies for manipulating major cellular systems through the utilization of cell cycle, apoptosis, immune modulation, epigenetic modification, and altered signal transduction pathways, including NF-kB, Notch, Wnt, MAPK, TLR, etc. We also discuss the important EBV latent antigens, namely EBNA1, EBNA2, EBNA3's and LMP's, which are important for targeting these major cellular pathways. KSHV infection progresses through the engagement of the activities of the major latent proteins LANA, v-FLIP and v-Cyclin, and the lytic replication and transcription activator (RTA). This review is a current, comprehensive approach that describes an in-depth understanding of gammaherpes viral encoded gene manipulation of the host system through targeting important biological processes in viral-associated cancers.

Dissecting the contribution of EBNA3C domains important for EBV-induced B-cell growth and proliferation.

Epstein-Barr virus (EBV) is an oncogenic gammaherpes virus which is linked to pathogenesis of several human lymphatic malignancies. The EBV essential latent antigen EBNA3C is critical for efficient conversion of primary human B-lymphocytes to lymphoblastic cell lines and for continued LCL growth. EBNA3C, an EBV latent antigen with oncogenic potential can bind and regulate the functions of a wide range of cellular transcription factors. In our current reverse genetics study, we deleted the full length EBNA3C, and independently the RBP-Jκ and Nm23-H1 binding sites within EBNA3C using BACmid recombinant engineering methodology. Our experiments demonstrated that deletion of the EBV EBNA3C open reading frame (ORF) and more specifically the residues 621-675 which binds Nm23H1 and SUMO-1 showed a significant reduction in the ability of the cells to proliferate. Furthermore, they exhibited lower infectivity of human peripheral blood mononuclear cells (PBMCs). We also showed that recombinant EBV with deletions of the EBNA3C ORF, as well as a recombinant with residues 621-675 within EBNA3C ORF deleted had diminished abilities to activate CD40. Our study also revealed that the full length (1-992) and 621-675 aa deletions of EBNA3C when compared to wild type EBV infected PBMCs had differential expression patterns for the phosphorylation of MAP kinases specifically p38, JNK and ERK. Regulation of ß-catenin also differed among wild type and EBNA3C deleted mutants. These temporal differences in signaling activities of these recombinant viruses in PBMCs is likely important in defining their functional importance in EBV-mediated B-cell transformation.

Regulation of the metastasis suppressor Nm23-H1 by tumor viruses.

Metastasis is the most common cause of cancer mortality. To increase the survival of patients, it is necessary to develop more effective methods for treating as well as preventing metastatic diseases. Recent advancement of knowledge in cancer metastasis provides the basis for development of targeted molecular therapeutics aimed at the tumor cell or its interaction with the host microenvironment. Metastasis suppressor genes (MSGs) are promising targets for inhibition of the metastasis process. During the past decade, functional significance of these genes, their regulatory pathways, and related downstream effector molecules have become a major focus of cancer research. Nm23-H1, first in the family of Nm23 human homologues, is a well-characterized, anti-metastatic factor linked with a large number of human malignancies. Mounting evidence to date suggests an important role for Nm23-H1 in reducing virus-induced tumor cell motility and migration. A detailed understanding of the molecular association between oncogenic viral antigens with Nm23-H1 may reveal the underlying mechanisms for tumor virus-associated malignancies. In this review, we will focus on the recent advances to our understanding of the molecular basis of oncogenic virus-induced progression of tumor metastasis by deregulation of Nm23-H1.

Kaposi's sarcoma-associated herpesvirus genome programming during the early stages of primary infection of peripheral blood mononuclear cells.

UNLABELLED: The early period of Kaposi's sarcoma-associated herpesvirus (KSHV) infection involves the dynamic expression of viral genes, which are temporally and epigenetically regulated. KSHV can effectively infect and persist in endothelial as well as human B cells with different gene expression patterns. To understand the temporal epigenetic changes which occur when KSHV infects the lymphocytic compartment, we infected human peripheral blood mononuclear cells (PBMCs) and comprehensively analyzed the changes which occurred at the binding sites of virally encoded lytic as well as latent proteins along with epigenetic modifications across the KSHV genome during early primary infection. Using chromatin immunoprecipitation (ChIP) assays, we showed that the KSHV genome acquires a uniquely distinct histone modification pattern of methylation (H3K4me3, H3K9me3, and H3K27me3) and acetylation (H3Ac) during de novo infection of human PBMCs. This pattern showed that the epigenetic changes were temporally controlled. The binding profiles of KSHV latent protein LANA and the immediate early proteins RTA and K8 showed specific patterns at different times postinfection, which reflects the gene expression program. Further analysis demonstrated that KSHV can concurrently express lytic and latent genes which were associated with histone modifications at these specific regions on the viral genome. We identified three KSHV genes, K3, ORF49, and ORF64, which exhibited different profiles of histone modifications during the early stages of PBMC infection. These studies established a distinct pattern of epigenetic modification which correlates with viral gene expression temporally regulated during the first 7 days of PBMC infection and provides clues to the regulatory program required for successful infection by KSHV of human PBMCs. IMPORTANCE: Kaposi's sarcoma-associated herpesvirus (KSHV) has been documented as one of the major contributors to morbidity and mortality in AIDS patients during the AIDS pandemic. During its life cycle, KSHV undergoes latent and lytic replication. Typically, KSHV maintains a stringent preference for latent infection in the infected B cells. However, 1 to 5% of infected cells undergo spontaneous lytic reactivation. KSHV lytic replication and infection of new cells are likely to be critical for maintaining the population of infected cells which drive virus-associated pathogenesis. Here, we explored the temporal changes of crucial histone marks on the KSHV genome during early infection of human primary peripheral blood mononuclear cells (PBMCs), which are a physiologically relevant system for monitoring primary infection. These results showed that KSHV possessed a distinct pattern of epigenetic marks during early infection of PBMCs. Further, KSHV concurrently expressed lytic and latent genes during this early period. These results now provide new evidence which contributes to understanding the molecular mechanism that regulates viral gene expression during early infection.

EBNA3C augments Pim-1 mediated phosphorylation and degradation of p21 to promote B-cell proliferation.

Epstein-Barr virus (EBV), a ubiquitous human herpesvirus, can latently infect the human population. EBV is associated with several types of malignancies originating from lymphoid and epithelial cell types. EBV latent antigen 3C (EBNA3C) is essential for EBV-induced immortalization of B-cells. The Moloney murine leukemia provirus integration site (PIM-1), which encodes an oncogenic serine/threonine kinase, is linked to several cellular functions involving cell survival, proliferation, differentiation, and apoptosis. Notably, enhanced expression of Pim-1 kinase is associated with numerous hematological and non-hematological malignancies. A higher expression level of Pim-1 kinase is associated with EBV infection, suggesting a crucial role for Pim-1 in EBV-induced tumorigenesis. We now demonstrate a molecular mechanism which reveals a direct role for EBNA3C in enhancing Pim-1 expression in EBV-infected primary B-cells. We also showed that EBNA3C is physically associated with Pim-1 through its amino-terminal domain, and also forms a molecular complex in B-cells. EBNA3C can stabilize Pim-1 through abrogation of the proteasome/Ubiquitin pathway. Our results demonstrate that EBNA3C enhances Pim-1 mediated phosphorylation of p21 at the Thr145 residue. EBNA3C also facilitated the nuclear localization of Pim-1, and promoted EBV transformed cell proliferation by altering Pim-1 mediated regulation of the activity of the cell-cycle inhibitor p21/WAF1. Our study demonstrated that EBNA3C significantly induces Pim-1 mediated proteosomal degradation of p21. A significant reduction in cell proliferation of EBV-transformed LCLs was observed upon stable knockdown of Pim-1. This study describes a critical role for the oncoprotein Pim-1 in EBV-mediated oncogenesis, as well as provides novel insights into oncogenic kinase-targeted therapeutic intervention of EBV-associated cancers.

Kaposi's sarcoma-associated herpesvirus-encoded LANA can induce chromosomal instability through targeted degradation of the mitotic checkpoint kinase Bub1.

UNLABELLED: Kaposi's sarcoma-associated herpesvirus (KSHV) has a significant contributory role in the development of three major human neoplastic or lymphoproliferative diseases: Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). These diseases are associated with chromosomal instability, a hallmark of human cancer. The latency-associated nuclear antigen (LANA) encoded by KSHV plays a key role in regulating a number of cellular pathways critical for oncogenesis. KSHV LANA alone can induce the development of B-cell hyperplasia and lymphoma in mice expressing LANA. LANA also induces chromosomal instability, thus promoting oncogenesis. However, the precise mechanism underlying LANA-mediated chromosomal instability remains uncharted. Here we report that LANA promoted the induction of chromosomal instability and the formation of micronuclei and multinucleation through its interaction with one of the critical spindle checkpoint proteins, Bub1, and the resulting degradation of Bub1. This interaction occurs through the Knl and kinase domains of Bub1, identified as important for stability and degradation. These results suggest that LANA can dysregulate Bub1 activity, which leads to aberrant chromosome replication and aneuploidy, thus contributing to KSHV-mediated oncogenesis. IMPORTANCE: This work represents the first set of results identifying a novel mechanism by which LANA, a latency-associated antigen encoded by KSHV, can induce the degradation of Bub1, a spindle checkpoint protein that is important for spindle checkpoint signaling and chromosome segregation. The downregulation of Bub1 mediated by LANA resulted in chromosomal instability, a hallmark of cancer. We further investigated the specific domains of Bub1 that are required for the interaction between LANA and Bub1. The results demonstrated that the Knl and kinase domains of Bub1 are required for the interaction between LANA and Bub1. In addition, we also investigated the mechanism by which LANA promoted Bub1 degradation. Our results showed that LANA interacted physically with the anaphase-promoting complex (APC/C), thus promoting the degradation of Bub1 in a ubiquitin-dependent process.

Inhibition of KAP1 enhances hypoxia-induced Kaposi's sarcoma-associated herpesvirus reactivation through RBP-Jκ.

UNLABELLED: Hypoxia-inducible factor 1α (HIF-1α) has been frequently implicated in many cancers as well as viral pathogenesis. Kaposi's sarcoma-associated herpesvirus (KSHV) is linked to several human malignancies. It can stabilize HIF-1α during latent infection and undergoes lytic replication in response to hypoxic stress. However, the mechanism by which KSHV controls its latent and lytic life cycle through the deregulation of HIF-1α is not fully understood. Our previous studies showed that the hypoxia-sensitive chromatin remodeler KAP1 was targeted by the KSHV-encoded latency-associated nuclear antigen (LANA) to repress expression of the major lytic replication and transcriptional activator (RTA). Here we further report that an RNA interference-based knockdown of KAP1 in KSHV-infected primary effusion lymphoma (PEL) cells disrupted viral episome stability and abrogated sub-G1/G1 arrest of the cell cycle while increasing the efficiency of KSHV lytic reactivation by hypoxia or using the chemical 12-O-tetradecanoylphorbol-13-acetate (TPA) or sodium butyrate (NaB). Moreover, KSHV genome-wide screening revealed that four hypoxia-responsive clusters have a high concurrence of both RBP-Jκ and HIF-1α binding sites (RBS+HRE) within the same gene promoter and are tightly associated with KAP1. Inhibition of KAP1 greatly enhanced the association of RBP-Jκ with the HIF-1α complex for driving RTA expression not only in normoxia but also in hypoxia. These results suggest that both KAP1 and the concurrence of RBS+HRE within the RTA promoter are essential for KSHV latency and hypoxia-induced lytic reactivation. IMPORTANCE: Kaposi's sarcoma-associated herpesvirus (KSHV), a DNA tumor virus, is an etiological agent linked to several human malignancies, including Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL). HIF-1α, a key hypoxia-inducible factor, is frequently elevated in KSHV latently infected tumor cells and contributes to KSHV lytic replication in hypoxia. The molecular mechanisms of how KSHV controls the latent and lytic life cycle through deregulating HIF-1α remain unclear. In this study, we found that inhibition of hypoxia-sensitive chromatin remodeler KAP1 in KSHV-infected PEL cells leads to a loss of viral genome and increases its sensitivity to hypoxic stress, leading to KSHV lytic reactivation. Importantly, we also found that four hypoxia-responsive clusters within the KSHV genome contain a high concurrence of RBP-Jκ (a key cellular regulator involved in Notch signaling) and HIF-1α binding sites. These sites are also tightly associated with KAP1. This discovery implies that KAP1, RBP-Jκ, and HIF-1α play an essential role in KSHV pathogenesis through subtle cross talk which is dependent on the oxygen levels in the infected cells.

Epstein-Barr virus essential antigen EBNA3C attenuates H2AX expression.

Epstein-Barr virus (EBV) latent antigen EBNA3C is implicated in B-cell immortalization and linked to several B-cell malignancies. Deregulation of H2AX can induce genomic instability with increased chromosomal aberrations, which ultimately leads to tumorigenesis. Here we demonstrated that EBNA3C can attenuate H2AX expression at the transcript and protein levels. A reduction of total H2AX levels was clearly observed upon infection of primary B cells with wild-type EBV but not with EBNA3C knockout recombinant EBV. H2AX also interacted with EBNA3C through its N-terminal domain (residues 1 to 100). Furthermore, H2AX mutated at Ser139 failed to interact with EBNA3C. Luciferase-based reporter assays also revealed that the binding domain of EBNA3C is sufficient for transcriptional inhibition of the H2AX promoter. EBNA3C also facilitated H2AX degradation through recruitment of components of the ubiquitin proteasome pathway. We further demonstrated that knockdown of H2AX in lymphoblastoid cell lines (LCLs) led to the upregulation of the Bub1 oncoprotein and downregulated expression of p53. Overall, our study provides additional insights into EBV-associated B-cell lymphomas, which are linked to the regulation of the DNA damage response system in infected cells. The importance of these insights are as follows: (i) EBNA3C downregulates H2AX expression at the protein and transcript levels in epithelial cells, B cells, and EBV-transformed LCLs, (ii) EBNA3C binds with wild-type H2AX but not with the Ser139 mutant of H2AX, (iii) the N terminus (residues 1 to 100) of EBNA3C is critical for binding to H2AX, (iv) localization of H2AX is predominantly nuclear in the presence of EBNA3C, and (v) H2AX knocked down in LCLs led to enhanced expression of Bub1 and downregulation of the tumor suppressor p53, which are both important for driving the oncogenic process.

Kaposi's sarcoma-associated herpesvirus-encoded LANA contributes to viral latent replication by activating phosphorylation of survivin.

UNLABELLED: Kaposi's sarcoma-associated herpesvirus (KSHV) is a human gammaherpesvirus casually linked to Kaposi's sarcoma (KS), multicentric Castleman's disease (MCD), and primary effusion lymphoma (PEL). Previously, we showed that LANA encoded by KSHV upregulates expression of survivin, a member of the inhibitor of apoptosis (IAP) family. This leads to an increase in the rate of cell proliferation of KSHV-infected B cells. LANA is required for tethering of the KSHV episome to the host chromosomes and efficiently segregates the viral genomes into dividing tumor cells. Here we show that LANA interacts with Aurora kinase B (AK-B) and induces phosphorylation of survivin at residue T34. Phosphorylation of survivin specifically on residue T34 enhances the activity of p300 and inhibits the activity of histone deacetylase 1 (HDAC-1), which then leads to an increase in acetylation of histone H3 on the viral genome. Phosphorylation of survivin specifically on residue T34 upregulates the activities of histone acetyltransferases and deacetylases, which then leads to an increase in viral copy number in KSHV-infected B cells. This results in a boost of KSHV replication in latently infected B-lymphoma cells. The studies showed that LANA can also function to regulate viral replication prior to mitosis of the latently infected cells, suggesting that LANA possesses a novel role in regulating KSHV replication in infected B cells. IMPORTANCE: This work represents a report of KSHV latent protein LANA and its interactions with AK-B leading to induction of phosphorylation of the oncoprotein survivin at residue T34. Phosphorylation of survivin specifically on residue T34 upregulates the activities of histone acetyltransferases and deacetylases. This leads to an increase in viral copy number in KSHV-infected B cells. These studies support a role for LANA in regulating KSHV replication through posttranslation modification in KSHV-infected B cells.

IRF-4-mediated CIITA transcription is blocked by KSHV encoded LANA to inhibit MHC II presentation.

Peptides presentation to T cells by MHC class II molecules is of importance in initiation of immune response to a pathogen. The level of MHC II expression directly influences T lymphocyte activation and is often targeted by various viruses. Kaposi's sarcoma-associated herpesvirus (KSHV) encoded LANA is known to evade MHC class I peptide processing, however, the effect of LANA on MHC class II remains unclear. Here, we report that LANA down-regulates MHC II expression and presentation by inhibiting the transcription of MHC II transactivator (CIITA) promoter pIII and pIV in a dose-dependent manner. Strikingly, although LANA knockdown efficiently disrupts the inhibition of CIITA transcripts from its pIII and pIV promoter region, the expression of HLA-DQß but no other MHC II molecules was significantly restored. Moreover, we revealed that the presentation of HLA-DQß enhanced by LANA knockdown did not help LANA-specific CD4+ T cell recognition of PEL cells, and the inhibition of CIITA by LANA is independent of IL-4 or IFN-γ signaling but dependent on the direct interaction of LANA with IRF-4 (an activator of both the pIII and pIV CIITA promoters). This interaction dramatically blocked the DNA-binding ability of IRF-4 on both pIII and pIV promoters. Thus, our data implies that LANA can evade MHC II presentation and suppress CIITA transcription to provide a unique strategy of KSHV escape from immune surveillance by cytotoxic T cells.