Aresté C, Blackbourn DJ (2006) HIV Tat-mediated transcriptional regulation of proteasome protein cleavage specificity., Biochem J 396 (2) pp. e13-e15
The major antigen-adapted immune response protecting a vertebrate against virus infection is that mediated by CTLs (cytotoxic T-lymphocytes). CTLs destroy virus-infected cells, thereby containing the infection. They are activated by recognition of peptide antigens or epitopes, presented to them in the context of MHC I proteins. These epitopes are derived from proteolytic degradation of endogenously synthesized proteins, which is mediated by the proteasome. Augmentation of epitope presentation by MHC I is thought to be effected by the immunoproteasome, induced in response to IFN-gamma (interferon-gamma) in some cells, and constitutively expressed in others. In this issue of the Biochemical Journal, Remoli and colleagues describe the manipulation of the immunoproteasome by the Tat (transcriptional activation) protein of HIV. The authors show that Tat deregulates the balance of the three proteins, LMP2 (low-molecular-mass polypeptide 2), LMP7 and MECL1 (multicatalytic endopeptidase complex-like 1), which distinguish the immunoproteasome from the proteasome, and they provide a molecular explanation. Intracellular Tat sequesters IRF-1 (interferon-regulatory factor-1) from its cognate promoter element, where normally it associates with STAT1 (signal transducer and activator of transcription 1) to activate basal transcription of the LMP2 gene. LMP2 expression is inhibited as a consequence, skewing the stoichiometry of the immunoproteasome and changing its enzymatic activity. These findings provide a molecular account of an immunomodulatory activity of HIV: changing the peptide antigen profile of cells expressing or exposed to Tat. They may also provide an avenue for manipulating vaccine efficacy and specificity with Tat-based adjuvants.
Colman R, Blackbourn DJ (2008) Risk factors in the development of Kaposi's sarcoma., AIDS 22 (13) pp. 1629-1632
Aresté C, Blackbourn DJ (2009) Modulation of the immune system by Kaposi's sarcoma-associated herpesvirus., Trends Microbiol 17 (3) pp. 119-129
The most recently identified human herpesvirus is Kaposi's sarcoma-associated herpesvirus (KSHV). It causes Kaposi's sarcoma, a tumour occurring most commonly in untreated AIDS patients and the leading cancer of men in certain parts of Africa. KSHV might also contribute to the pathogenesis of primary effusion lymphoma and multicentric Castleman's disease. The genome of KSHV contains 86 genes, almost a quarter of which encode proteins with either demonstrated or potential immunoregulatory activity. They include homologues of cellular proteins and unique KSHV proteins that can deregulate many aspects of the immune response, including T- and B-cell functions, complement activation, the innate antiviral interferon response and natural killer cell activity. The functions of these proteins and the ways in which they perturb the normal immune response are the subjects of the present review.
Amini AA, Solovyova AS, Sadeghian H, Blackbourn DJ, Rezaee SA (2014) Structural properties of a viral orthologue of cellular CD200 protein: KSHV vOX2., Virology 474 pp. 94-104
Kaposiós sarcoma-associated herpesvirus (KSHV) vOX2 is a cell surface glycoprotein expressed during viral lytic replication to suppress host inflammatory reactions. Here we have characterised vOX2 with biochemical, biophysical and bioinformatics tools and as a result propose a 3-dimensional model for vOX2 based on structural and functional homology with the PD-L1 protein. To validate this model, vOX2 was characterised by analytical ultracentrifugation (AUC) and circular dichroism spectroscopy (CD). The results identified the potential glycosylation sites and revealed that vOX2 is predominantly a beta-folded molecule with an RGD adhesion motif exposed on the C-terminal domain. The protein exists in monomer-dimer equilibrium similar to its IgV-type folded homologues, with 30-36% glycosylation and the molecular weight of the extracellular fragment of molecule is 32.0-33.6 kDa, much less than 50 kDa. Thus, the structural similarity to PD-L1 verifies its immunomodulatory potential and the RGD motif suggests an adhesive capacity.
McKimmie CS, Singh MD, Hewit K, Lopez-Franco O, Le Brocq M, Rose-John S, Lee KM, Baker AH, Wheat R, Blackbourn DJ, Nibbs RJB, Graham GJ (2013) An analysis of the function and expression of D6 on lymphatic endothelial cells, BLOOD 121 (18) pp. 3768-3777 AMER SOC HEMATOLOGY
Jeffery HC, Wheat RL, Blackbourn DJ, Nash GB, Butler LM (2013) Infection and transmission dynamics of rKSHV.219 in primary endothelial cells., J Virol Methods 193 (1) pp. 251-259
Kaposi's sarcoma-associated herpesvirus (KSHV) is the aetiologic agent of Kaposi's sarcoma (KS), a tumour of endothelial cell origin. The study of KS development was aided by the generation of a recombinant GFP (latent)/RFP (lytic)-expressing KSHV (rKSHV.219) by Vieira and O'Hearn (2004). In this study the first data characterising primary endothelial cell infection and transmission with this virus is presented. Infection was predominantly latent and the percentage of GFP-positive cells increased over time. Neither horizontal transmission of infection, nor cellular proliferation, explained this increase. Analysis of latency-associated nuclear antigen (LANA-1) expression revealed that a threshold level of infection was required for GFP expression early post infection. At later time points GFP correlated more closely with LANA-1 expression, likely due to the accumulation of GFP over time. This study provides methodological guidance for the use of rKSHV.21. In addition, it highlights potential problems associated with the use of fluorescent proteins as markers of viral infection.
Butler LM, Wheat RL, Jeffery HC, Nash GB, Blackbourn DJ (2009) Infection of endothelial cells with Kaposi's sarcoma-associated herpesvirus selectively inhibits recruitment of flowing neutrophils in an inflammatory model, CYTOKINE 48 (1-2) pp. 51-51 ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
Butler LM, Jeffery HC, Wheat RL, Long HM, Rae PC, Nash GB, Blackbourn DJ (2012) Kaposi's sarcoma-associated herpesvirus inhibits expression and function of endothelial cell major histocompatibility complex class II via suppressor of cytokine signaling 3., J Virol 86 (13) pp. 7158-7166
Endothelial cells (EC) can present antigen to either CD8(+) T lymphocytes through constitutively expressed major histocompatibility complex class I (MHC-I) or CD4(+) T lymphocytes through gamma interferon (IFN-³)-induced MHC-II. Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of Kaposi's sarcoma (KS), an EC neoplasm characterized by dysregulated angiogenesis and a substantial inflammatory infiltrate. KSHV is understood to have evolved strategies to inhibit MHC-I expression on EC and MHC-II expression on primary effusion lymphoma cells, but its effects on EC MHC-II expression are unknown. Here, we report that the KSHV infection of human primary EC inhibits IFN-³-induced expression of the MHC-II molecule HLA-DR at the transcriptional level. The effect is functionally significant, since recognition by an HLA-DR-restricted CD4(+) T-cell clone in response to cognate antigen presented by KSHV-infected EC was attenuated. Inhibition of HLA-DR expression was also achieved by exposing EC to supernatant from KSHV-inoculated EC before IFN-³ treatment, revealing a role for soluble mediators. IFN-³-induced phosphorylation of STAT-1 and transcription of CIITA were suppressed in KSHV-inoculated EC via a mechanism involving SOCS3 (suppressor of cytokine signaling 3). Thus, KSHV infection resulted in transcriptional upregulation of SOCS3, and treatment with RNA interference against SOCS3 relieved virus-induced inhibition of IFN-³-induced STAT-1 phosphorylation. Since cell surface MHC-II molecules present peptide antigens to CD4(+) T lymphocytes that can function either as direct cytolytic effectors or to initiate and regulate adaptive immune responses, inhibition of this antigen-presenting pathway would provide a survival advantage to the virus.
Wheat R, Roberts C, Waterboer T, Steele J, Marsden J, Steven NM, Blackbourn DJ (2014) Inflammatory cell distribution in primary Merkel cell carcinoma, Cancers 6 (2) pp. 1047-1064
Merkel cell carcinoma (MCC) is an aggressive poorly differentiated neuroendocrine cutaneous carcinoma associated with older age, immunodeficiency and Merkel cell polyomavirus (MCPyV) integrated within malignant cells. The presence of intra-tumoural CD8 lymphocytes reportedly predicts better MCC-specific survival. In this study, the distribution of inflammatory cells and properties of CD8+ T lymphocytes within 20 primary MCC specimens were characterised using immunohistochemistry and multicolour immunofluorescent staining coupled to confocal microscopy. CD8+ cells and CD68+ macrophages were identified in 19/20 primary MCC. CD20+ B cells were present in 5/10, CD4+ cells in 10/10 and FoxP3+ cells in 7/10 specimens. Only two specimens had almost no inflammatory cells. Within specimens, inflammatory cells followed the same patchy distribution, focused at the edge of sheets and nodules and, in some cases, more intense in trabecular areas. CD8+ cells were outside vessels on the edge of tumour. Those few within malignant sheets typically lined up in fine septa not contacting MCC cells expressing MCPyV large T antigen. The homeostatic chemokine CXCL12 was expressed outside malignant nodules whereas its receptor CXCR4 was identified within tumour but not on CD8+ cells. CD8+ cells lacked CXCR3 and granzyme B expression irrespective of location within stroma versus malignant nodules or of the intensity of the intra-tumoural infiltrate. In summary, diverse inflammatory cells were organised around the margin of malignant deposits suggesting response to aberrant signaling, but were unable to penetrate the tumour microenvironment itself to enable an immune response against malignant cells or their polyomavirus. © 2014 by the authors; licensee MDPI, Basel, Switzerland.
Wang L, Pietrek M, Brinkmann MM, Haevemeier A, Fischer I, Hillenbrand B, Dittrich-Breiholz O, Kracht M, Chanas S, Blackbourn DJ, Schulz TF (2009) Identification and functional characterization of a spliced rhesus rhadinovirus gene with homology to the K15 gene of Kaposi's sarcoma-associated herpesvirus, JOURNAL OF GENERAL VIROLOGY 90 pp. 1190-1201 SOC GENERAL MICROBIOLOGY
Okroj M, Mark L, Stokowska A, Wong SW, Rose N, Blackbourn DJ, Villoutreix BO, Spiller OB, Blom AM (2009) Characterization of the Complement Inhibitory Function of Rhesus Rhadinovirus Complement Control Protein (RCP), JOURNAL OF BIOLOGICAL CHEMISTRY 284 (1) pp. 505-514 AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Mark L, Proctor DG, Blackbourn DJ, Blom AM, Spiller OB (2008) Separation of decay-accelerating and cofactor functional activities of Kaposi's sarcoma-associated herpesvirus complement control protein using monoclonal antibodies, IMMUNOLOGY 123 (2) pp. 228-238 BLACKWELL PUBLISHING
Ye F-C, Blackbourn DJ, Mengel M, Xie J-P, Qian L-W, Greene W, Yeh I-T, Graham D, Gao S-J (2007) Kaposi's sarcoma-associated herpesvirus promotes angiogenesis by inducing angiopoietin-2 expression via AP-1 and Ets1, JOURNAL OF VIROLOGY 81 (8) pp. 3980-3991 AMER SOC MICROBIOLOGY
Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic virus, the etiological agent of Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL). One of the key viral proteins that contributes to tumorigenesis is vFLIP, a viral homolog of the FLICE inhibitory protein. This KSHV protein interacts with the NFºB pathway to trigger the expression of antiapoptotic and proinflammatory genes and ultimately leads to tumor formation. The expression of vFLIP is regulated at the translational level by an internal ribosomal entry site (IRES) element. However, the precise mechanism by which ribosomes are recruited internally and the exact location of the IRES has remained elusive. Here we show that a 252-nt fragment directly upstream of vFLIP, within a coding region, directs translation. We have established its RNA structure and demonstrate that IRES activity requires the presence of eIF4A and an intact eIF4G. Furthermore, and unusually for an IRES, eIF4E is part of the complex assembled onto the vFLIP IRES to direct translation. These molecular interactions define a new paradigm for IRES-mediated translation.
Hollingworth R, Skalka GL, Stewart GS, Hislop AD, Blackbourn DJ, Grand RJ (2015) Activation of DNA Damage Response Pathways during Lytic Replication of KSHV., Viruses 7 (6) pp. 2908-2927
Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of several human malignancies. Human tumour viruses such as KSHV are known to interact with the DNA damage response (DDR), the molecular pathways that recognise and repair lesions in cellular DNA. Here it is demonstrated that lytic reactivation of KSHV leads to activation of the ATM and DNA-PK DDR kinases resulting in phosphorylation of multiple downstream substrates. Inhibition of ATM results in the reduction of overall levels of viral replication while inhibition of DNA-PK increases activation of ATM and leads to earlier viral release. There is no activation of the ATR-CHK1 pathway following lytic replication and CHK1 phosphorylation is inhibited at later times during the lytic cycle. Despite evidence of double-strand breaks and phosphorylation of H2AX, 53BP1 foci are not consistently observed in cells containing lytic virus although RPA32 and MRE11 localise to sites of viral DNA synthesis. Activation of the DDR following KSHV lytic reactivation does not result in a G1 cell cycle block and cells are able to proceed to S-phase during the lytic cycle. KSHV appears then to selectively activate DDR pathways, modulate cell cycle progression and recruit DDR proteins to sites of viral replication during the lytic cycle.
Jackson BR, Boyne JR, Noerenberg M, Taylor A, Hautbergue GM, Walsh MJ, Wheat R, Blackbourn DJ, Wilson SA, Whitehouse A (2011) An Interaction between KSHV ORF57 and UIF Provides mRNA-Adaptor Redundancy in Herpesvirus Intronless mRNA Export, PLOS PATHOGENS 7 (7) ARTN e1002138 PUBLIC LIBRARY SCIENCE
Knight LM, Stakaityte G, Wood JJ, Abdul-Sada H, Griffiths DA, Howell GJ, Wheat R, Blair GE, Steven NM, Macdonald A, Blackbourn DJ, Whitehouse A (2014) Merkel cell polyomavirus small T antigen mediates microtubule destabilization to promote cell motility and migration., J Virol 89 (1) pp. 35-47
UNLABELLED: Merkel cell carcinoma (MCC) is an aggressive skin cancer of neuroendocrine origin with a high propensity for recurrence and metastasis. Merkel cell polyomavirus (MCPyV) causes the majority of MCC cases due to the expression of the MCPyV small and large tumor antigens (ST and LT, respectively). Although a number of molecular mechanisms have been attributed to MCPyV tumor antigen-mediated cellular transformation or replication, to date, no studies have investigated any potential link between MCPyV T antigen expression and the highly metastatic nature of MCC. Here we use a quantitative proteomic approach to show that MCPyV ST promotes differential expression of cellular proteins implicated in microtubule-associated cytoskeletal organization and dynamics. Intriguingly, we demonstrate that MCPyV ST expression promotes microtubule destabilization, leading to a motile and migratory phenotype. We further highlight the essential role of the microtubule-associated protein stathmin in MCPyV ST-mediated microtubule destabilization and cell motility and implicate the cellular phosphatase catalytic subunit protein phosphatase 4C (PP4C) in the regulation of this process. These findings suggest a possible molecular mechanism for the highly metastatic phenotype associated with MCC. IMPORTANCE: Merkel cell polyomavirus (MCPyV) causes the majority of cases of Merkel cell carcinoma (MCC), an aggressive skin cancer with a high metastatic potential. However, the molecular mechanisms leading to virally induced cancer development have yet to be fully elucidated. In particular, no studies have investigated any potential link between the virus and the highly metastatic nature of MCC. We demonstrate that the MCPyV small tumor antigen (ST) promotes the destabilization of the host cell microtubule network, which leads to a more motile and migratory cell phenotype. We further show that MCPyV ST induces this process by regulating the phosphorylation status of the cellular microtubule-associated protein stathmin by its known association with the cellular phosphatase catalytic subunit PP4C. These findings highlight stathmin as a possible biomarker of MCC and as a target for novel antitumoral therapies.
Rezaee SA, Cunningham C, Davison AJ, Blackbourn DJ (2006) Kaposi's sarcoma-associated herpesvirus immune modulation: an overview., J Gen Virol 87 (Pt 7) pp. 1781-1804
Kaposi's sarcoma-associated herpesvirus (KSHV) is the most recently discovered human herpesvirus. It is the aetiological agent of Kaposi's sarcoma (KS), a tumour frequently affecting AIDS patients not receiving treatment. KSHV is also a likely cause of two lymphoproliferative diseases: multicentric Castleman's disease and primary effusion lymphoma. The study of KSHV offers exciting challenges for understanding the mechanisms of virus pathogenesis, including those involved in establishing infection and dissemination in the host. To facilitate these processes, approximately one-quarter of KSHV genes encode cellular homologues or unique proteins that have immunomodulatory roles in cytokine production, apoptosis, cell signalling and the immunological synapse. The activities of these molecules are considered in the present review and the positions of their genes are mapped from a complete KSHV genome sequence derived from a KS biopsy. The understanding gained enables the significance of different components of the immune response in protection against KSHV infection to be evaluated. It also helps to unravel the complexities of cellular and immunological pathways and offers the potential for exploiting viral immunomodulators and derivatives in disease therapy.
Mutocheluh M, Hindle L, Aresté C, Chanas SA, Butler LM, Lowry K, Shah K, Evans DJ, Blackbourn DJ (2011) Kaposi's sarcoma-associated herpesvirus viral interferon regulatory factor-2 inhibits type 1 interferon signalling by targeting interferon-stimulated gene factor-3., J Gen Virol 92 (Pt 10) pp. 2394-2398
Kaposi's sarcoma-associated herpesvirus (KSHV) encodes four viral interferon regulatory factors (vIRF-1-4). We investigated the mechanism and consequences of vIRF-2-mediated inhibition of interferon-response element signalling following type I interferon (IFN) induction. Western blot and electrophoretic mobility-shift assays identified the interferon-stimulated gene factor-3 (ISGF-3) components STAT1 and IRF-9 as the proximal targets of vIRF-2 activity. The biological significance of vIRF-2 inhibition of ISGF-3 was demonstrated by vIRF-2-mediated rescue of the replication of the IFN-sensitive virus encephalomyocarditis virus. This study provides both a mechanism and evidence for KSHV vIRF-2-mediated suppression of the consequences of type 1 IFN-induced signalling.
Misstear K, Chanas SA, Rezaee SA, Colman R, Quinn LL, Long HM, Goodyear O, Lord JM, Hislop AD, Blackbourn DJ (2012) Suppression of antigen-specific T cell responses by the Kaposi's sarcoma-associated herpesvirus viral OX2 protein and its cellular orthologue, CD200., J Virol 86 (11) pp. 6246-6257
Regulating appropriate activation of the immune response in the healthy host despite continual immune surveillance dictates that immune responses must be either self-limiting and therefore negatively regulated following their activation or prevented from developing inappropriately. In the case of antigen-specific T cells, their response is attenuated by several mechanisms, including ligation of CTLA-4 and PD-1. Through the study of the viral OX2 (vOX2) immunoregulator encoded by Kaposi's sarcoma-associated herpesvirus (KSHV), we have identified a T cell-attenuating role both for this protein and for CD200, a cellular orthologue of the viral vOX2 protein. In vitro, antigen-presenting cells (APC) expressing either native vOX2 or CD200 suppressed two functions of cognate antigen-specific T cell clones: gamma interferon (IFN-³) production and mobilization of CD107a, a cytolytic granule component and measure of target cell killing ability. Mechanistically, vOX2 and CD200 expression on APC suppressed the phosphorylation of ERK1/2 mitogen-activated protein kinase in responding T cells. These data provide the first evidence for a role of both KSHV vOX2 and cellular CD200 in the negative regulation of antigen-specific T cell responses. They suggest that KSHV has evolved to harness the host CD200-based mechanism of attenuation of T cell responses to facilitate virus persistence and dissemination within the infected individual. Moreover, our studies define a new paradigm in immune modulation by viruses: the provision of a negative costimulatory signal to T cells by a virus-encoded orthologue of CD200.
Taylor GS, Blackbourn DJ (2011) Infectious agents in human cancers: lessons in immunity and immunomodulation from gammaherpesviruses EBV and KSHV., Cancer Lett 305 (2) pp. 263-278
Members of the herpesvirus family have evolved the ability to persist in their hosts by establishing a reservoir of latently infected cells each carrying the viral genome with reduced levels of viral protein synthesis. In order to spread within and between hosts, in some cells, the quiescent virus will reactivate and enter lytic cycle replication to generate and release new infectious virus particles. To allow the efficient generation of progeny viruses, all herpesviruses have evolved a wide variety of immunomodulatory mechanisms to limit the exposure of cells undergoing lytic cycle replication to the immune system. Here we have focused on the human gammaherpesviruses Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV) that, uniquely among the eight human herpesviruses identified to date, have growth transforming potential. Most people infected with these viruses will not develop cancer, viral growth-transforming activity being kept under control by the host's antigen-specific immune responses. Nonetheless, EBV and KSHV are associated with several malignancies in which various viral proteins, either predominantly or exclusively latency-associated, are expressed; at least some of these proteins also have immunomodulatory activities. Of these malignancies, some are the result of a disrupted virus/immune balance through genetic, infectious or iatrogenic immune suppression. Others develop in people that are not overtly immune suppressed and likely modulate the immunological response. This latter aspect of immune modulation by EBV and KSHV forms the basis of this review.
Fuld S, Cunningham C, Klucher K, Davison AJ, Blackbourn DJ (2006) Inhibition of interferon signaling by the Kaposi's sarcoma-associated herpesvirus full-length viral interferon regulatory factor 2 protein, JOURNAL OF VIROLOGY 80 (6) pp. 3092-3097 AMER SOC MICROBIOLOGY
Okroj M, Mark L, Stokowska A, Wong S, Rose N, Blackbourn D, Villoutreix B, Spiller B, Blom A (2008) Characterization of the complement inhibitory function of Rhesus rhadinovirus, MOLECULAR IMMUNOLOGY 45 (16) pp. 4172-4173 PERGAMON-ELSEVIER SCIENCE LTD
Butler LM, Jeffery HC, Wheat RL, Rae PC, Townsend K, Alkharsah KR, Schulz TF, Nash GB, Blackbourn DJ (2011) Kaposi's sarcoma-associated herpesvirus infection of endothelial cells inhibits neutrophil recruitment through an interleukin-6-dependent mechanism: a new paradigm for viral immune evasion., J Virol 85 (14) pp. 7321-7332
Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of Kaposi's sarcoma (KS), an endothelial cell (EC) neoplasm characterized by dysregulated angiogenesis and inflammation. KSHV infection of EC causes production of proinflammatory mediators, regarded as possible initiators of the substantial mononuclear leukocyte recruitment seen in KS. Conversely, KSHV immune evasion strategies exist, such as degradation of EC leukocyte adhesion receptors by viral proteins. Here, we report the effects of KSHV infection of primary EC on recruitment of flowing leukocytes. Infection did not initiate adhesion of any leukocyte subset per se. However, on cytokine-stimulated EC, KSHV specifically inhibited neutrophil, but not PBL or monocyte, transmigration, an observation consistent with the inflammatory cell profile found in KS lesions in vivo. This inhibition could be recapitulated on uninfected EC using supernatant from infected cultures. These supernatants contained elevated levels of human interleukin 6 (hIL-6), and both the KSHV- and the supernatant-induced inhibitions of neutrophil transmigration were abrogated in the presence of a hIL-6 neutralizing antibody. Furthermore, preconditioning of EC with hIL-6 mimicked the effect of KSHV. Using RNA interference (RNAi), we show that upregulation of suppressor of cytokine signaling 3 (SOCS3) was necessary for this effect of hIL-6. These studies reveal a novel paracrine mode of KSHV immune evasion, resulting in reduced recruitment of neutrophils, a cell type whose antiviral and antitumor roles are becoming increasingly appreciated. Moreover, the findings have implications for our understanding of the contribution of hIL-6 to the pathogenesis of other inflammatory disorders and tumors in which this cytokine is abundant.
Aresté C, Mutocheluh M, Blackbourn DJ (2009) Identification of caspase-mediated decay of interferon regulatory factor-3, exploited by a Kaposi sarcoma-associated herpesvirus immunoregulatory protein., J Biol Chem 284 (35) pp. 23272-23285
Upon virus infection, the cell mounts an innate type I interferon (IFN) response to limit the spread. This response is orchestrated by the constitutively expressed IFN regulatory factor (IRF)-3 protein, which becomes post-translationally activated. Although the activation events are understood in detail, the negative regulation of this innate response is less well understood. Many viruses, including Kaposi sarcoma-associated herpesvirus (KSHV), have evolved defense strategies against this IFN response. Thus, KSHV encodes a viral IRF (vIRF)-2 protein, sharing homology with cellular IRFs and is a known inhibitor of the innate IFN response. Here, we show that vIRF-2 mediates IRF-3 inactivation by a mechanism involving caspase-3, although vIRF-2 itself is not pro-apoptotic. Importantly, we also show that caspase-3 participates in normal IRF-3 turnover in the absence of vIRF-2, during the antiviral response induced by poly(I:C) transfection. These data provide unprecedented insight into negative regulation of IRF-3 following activation of the type I IFN antiviral response and the mechanism by which KSHV vIRF-2 inhibits this innate response.
Mark L, Spiller OB, Blackbourn DJ, Blom AM (2007) Molecular details of the complement regulatory and cell attaching functions of KCP, MOLECULAR IMMUNOLOGY 44 (1-3) pp. 213-213 PERGAMON-ELSEVIER SCIENCE LTD
Durrington HJ, Upton PD, Hoer S, Boname J, Dunmore BJ, Yang J, Crilley TK, Butler LM, Blackbourn DJ, Nash GB, Lehner PJ, Morrell NW (2010) Identification of a Lysosomal Pathway Regulating Degradation of the Bone Morphogenetic Protein Receptor Type II, JOURNAL OF BIOLOGICAL CHEMISTRY 285 (48) pp. 37641-37649 AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Damania B, Blackbourn DJ (2012) Innate barriers to viral infection, Future Microbiology 7 (7) pp. 815-822
Innate immunity represents the foremost barrier to viral infection. In order to infect a cell efficiently, viruses need to evade innate immune effectors such as interferons and inflammatory cytokines. Pattern recognition receptors can detect viral components or pathogen-associated molecular patterns. These receptors then elicit innate immune responses that result in the generation of type I interferons and proinflammatory cytokines. Organized by the Society for General Microbiology, one session of this conference focused on the current state-of-the-art knowledge on innate barriers to infection of different RNA and DNA viruses. Experts working on innate immunity in the context of viral infection provided insight into different aspects of innate immune recognition and also discussed areas for future research. Here, we provide an overview of the session on innate barriers to infection. © 2012 Future Medicine Ltd.
Jacobs SR, Gregory SM, West JA, Wollish AC, Bennett CL, Blackbourn DJ, Heise MT, Damania B (2013) The Viral Interferon Regulatory Factors of Kaposi's Sarcoma-Associated Herpesvirus Differ in Their Inhibition of Interferon Activation Mediated by Toll-Like Receptor 3, JOURNAL OF VIROLOGY 87 (2) pp. 798-806 AMER SOC MICROBIOLOGY
Berhane S, Aresté C, Ablack JN, Ryan GB, Blackbourn DJ, Mymryk JS, Turnell AS, Steele JC, Grand RJ (2011) Adenovirus E1A interacts directly with, and regulates the level of expression of, the immunoproteasome component MECL1., Virology 421 (2) pp. 149-158
Proteasomes represent the major non-lysosomal mechanism responsible for the degradation of proteins. Following interferon ³ treatment 3 proteasome subunits are replaced producing immunoproteasomes. Adenovirus E1A interacts with components of the 20S and 26S proteasome and can affect presentation of peptides. In light of these observations we investigated the relationship of AdE1A to the immunoproteasome. AdE1A interacts with the immunoproteasome subunit, MECL1. In contrast, AdE1A binds poorly to the proteasome ²2 subunit which is replaced by MECL1 in the conversion of proteasomes to immunoproteasomes. Binding sites on E1A for MECL1 correspond to the N-terminal region and conserved region 3. Furthermore, AdE1A causes down-regulation of MECL1 expression, as well as LMP2 and LMP7, induced by interferon ³ treatment during Ad infections or following transient transfection. Consistent with previous reports AdE1A reduced IFN³-stimulated STAT1 phosphorylation which appeared to be responsible for its ability to reduce expression of immunoproteasome subunits.
Stakaityt? Gabriel?, Nwogu Nnenna, Dobson Samuel J., Knight Laura M., Wasson Christopher W., Salguero Bodes Francisco, Blackbourn David, Blair G. Eric, Mankouri Jamel, Macdonald Andrew, Whitehouse Adrian (2017) Merkel cell polyomavirus small T antigen drives cell motility via Rho-GTPase-induced filopodia formation, Journal of Virology 92 (2) e00940-17 pp. 1-21
American Society for Microbiology
Cell motility and migration is a complex, multi-step, and multi-component process, intrinsic to progression and metastasis. Motility is dependent on the activity of integrin receptors and Rho-family GTPases resulting in the remodelling of the actin cytoskeleton and formation of various motile actin-based protrusions. Merkel cell carcinoma (MCC) is an aggressive skin cancer with a high likelihood of recurrence and metastasis. Merkel cell polyomavirus (MCPyV) is associated with the majority of MCC cases, and MCPyV-induced tumourigenesis largely depends on the expression of the small tumour antigen (ST). Since the discovery of MCPyV, a number of mechanisms have been suggested to account for replication and tumourigenesis, but to date, little is known about potential links between MCPyV T antigen expression and the metastatic nature of MCC. Previously, we have described the action of MCPyV ST on the microtubule network and how this impacts on cell motility and migration. Here we demonstrate that MCPyV ST affects the actin cytoskeleton, to promote the formation of filopodia, through a mechanism involving the catalytic subunit of protein phosphatase 4 (PP4C). We also show that MCPyV ST-induced cell motility is dependent upon the activity of Rho-family GTPases Cdc42 and RhoA. In addition, our results indicate that the MCPyV ST-PP4C interaction results in the dephosphorylation of ²1 integrin, likely driving the cell motility pathway. These findings describe a novel mechanism by which a tumour virus induces cell motility, which may ultimately lead to cancer metastasis and provides opportunities and strategies for targeted interventions for disseminated MCC.
Aflatoxin B1 (AFB1) contamination of food is very high in most sub-Saharan African countries. AFB1 is known to cause hepatocellular carcinoma (HCC) by inducing mutation in the tumour suppressor gene TP53. The number of new HCC cases is high in West Africa with an accompanying high mortality. The type I interferon (IFN) pathway of the innate immune system limits viral infections and exerts its anti-cancer property by up-regulating tumour suppressor activities and pro-apoptotic pathways. Indeed, IFN-± is reported to show significant protective effects against hepatic fibrogenesis and carcinogenesis. However, the mechanism behind AFB1 deregulation of the type I interferon (IFN) signalling pathway, with consequent HCC is largely unknown. This current study seeks to test the hypothesis that AFB1 inhibits the type I IFN response by directly interfering with key signalling proteins and thus increase the risk of HCC in humans.
We evaluated the effects of AFB1 on the type I IFN signalling pathway using IFN stimulated response element (ISRE)-based luciferase reporter gene assay. In addition, the effects of AFB1 on the transcript levels of JAK1, STAT1 and OAS3 were assessed by real-time quantitative polymerase chain reaction (RT-qPCR) and confirmed by immunoblot assay.
Our results indicated that AFB1 inhibited the type I IFN signalling pathway in human hepatoma cell line HepG2 cells by suppressing the transcript levels of JAK1, STAT1 and OAS3. AFB1 also decreased the accumulation of STAT1 protein.
The inhibition of the type I IFN anti-cancer response pathway by AFB1 suggest a novel mechanism by which AFB1 may induce hepatocellular carcinoma in humans.
McHugh Donal, Caduff Nicole, Barros Mario Henrique M., Rämer Patrick C., Raykova Ana, Murer Anita, Landtwing Vanessa, Quast Isaak, Styles Christine T., Spohn Michael, Fowotade Adeola, Delecluse Henri-Jacques, Papoudou-Bai Alexandra, Lee Yong-Moon, Kim Jin-Man, Middeldorp Jaap, Schulz Thomas F., Cesarman Ethel, Zbinden Andrea, Capaul Riccarda, White Robert E., Allday Martin J., Niedobitek Gerald, Blackbourn David, Grundhoff Adam, Münz Christian (2017) Persistent KSHV Infection Increases EBV-Associated Tumor Formation In Vivo via Enhanced EBV Lytic Gene Expression, Cell Host & Microbe 22 (1) pp. 61-73.e7
The human tumor viruses Epstein-Barr virus (EBV) and Kaposi sarcoma-associated herpesvirus (KSHV) establish persistent infections in B cells. KSHV is linked to primary effusion lymphoma (PEL), and 90% of PELs also contain EBV. Studies on persistent KSHV infection in vivo and the role of EBV co-infection in PEL development have been hampered by the absence of small animal models. We developed mice reconstituted with human immune system components as a model for KSHV infection and find that EBV/KSHV dual infection enhanced KSHV persistence and tumorigenesis. Dual-infected cells displayed a plasma cell-like gene expression pattern similar to PELs. KSHV persisted in EBV-transformed B cells and was associated with lytic EBV gene expression, resulting in increased tumor formation. Evidence of elevated lytic EBV replication was also found in EBV/KSHV dually infected lymphoproliferative disorders in humans. Our data suggest that KSHV augments EBV-associated tumorigenesis via stimulation of lytic EBV replication.
Objective: Vitamin D deficiency (serum 25-hydroxyvitamin DÂ25nmol/L) is extremely common in western-dwelling South Asians but evidence regarding vitamin D supplement usage in this group is very limited. This work identifies demographic, dietary and lifestyle predictors associated with vitamin D supplement use.
Design: Cross-sectional analysis of baseline vitamin D supplement use data.
Setting: UK Biobank cohort.
Subjects: In total, n 8024 South Asians (Bangladeshi, Indian, Pakistani), aged 40-69 years.
Results: Twenty-three % of men and 39% of women (PÂ0.001) [22% of Bangladeshis, 32% of Indians, 25% of Pakistanis (PÂ0.001)] took a vitamin D containing supplement. Median vitamin D intakes from diet were low at 1.0-3.0 micrograms per day, being highest in Bangladeshis and lowest in Indians (PÂ0.001). Logistic regression modelling showed that females had a higher odds of vitamin D supplement use than males (odds ratio (OR) = 2.02; 95% confidence interval (CI) 1.79 to 2.28). A lower supplement usage was seen in younger persons (40-60 years) (OR=0.75; 95% CI 0.65 to 0.86 reference= Ã60 years), and those living outside of Greater London (OR=0.53 to 0.77), with borderline trends for a lower body mass index, higher oily fish intake and higher household income associated with increased odds of vitamin D supplement use.
Conclusions: Vitamin D supplements were not used by most South Asians and intakes from diet alone are likely to be insufficient to maintain adequate vitamin D status. Public health strategies are now urgently required to promote the use of vitamin D supplements in these specific UK South Asian sub-groups.