The Roman centuriated landscape: conception, genesis and development as inferred from the Ager Tarraconensis case

Although centuriation was only one system of Roman land division, its impact on the landscape and its visibility in modern field arrangements make it the most commonly recognized expression of Roman landscapes. Centuriated grid systems are usually analyzed from a materialistic point of view and consequently regarded as an assertion of Roman dominance over conquered territories. In this sense, their productive function is clear. The hinterland of Tarraco (the ancient capital of the Roman province of Tarraconensis) offers one of the most clearly documented examples of multiple-grid centuriated systems. From 2006 to 2010, the Landscape Archaeology Research Group (GIAP) of the Catalan Institute of Classical Archaeology employed a wide array of digital and field methodologies at Tarraco to record the traces of centuriated land divisions and their Roman origin. Most importantly, these methods have allowed research to move beyond pure description of the traces to explore the concepts and ideas behind the making of a centuriated landscape. By using Tarraco as a case study, this article shows how centuriation was not only a system for dividing the land but also a conceptual appropriation of the landscape based on a strong mythical and religious backgroun

The antiviral protein viperin inhibits HCV replication via interaction with NS5A

The interferon-stimulated gene viperin has been shown to have antiviral activity against hepatitis C virus (HCV) in the context of the HCV replicon, although the molecular mechanisms responsible are not well understood. Here we demonstrate that viperin plays an integral part in the ability of interferon to limit replication of cell culture derived HCV (JFH-1) that accurately reflects the complete viral life cycle. Using confocal microscopy and Fluorescence Resonance Energy Transfer (FRET) analysis we demonstrate that viperin localizes and interacts with HCV NS5A at the lipid droplet interface. In addition viperin also associates with NS5A and the pro-viral cellular factor, VAP-A at the HCV replication complex. The ability of viperin to limit HCV replication was dependent on residues within the C-terminus as well as an N-terminal amphipathic helix. Removal of the amphipathic helix redirected viperin from the cytosolic face of the ER and the lipid droplet to a homogenous cytoplasmic distribution, coinciding with a loss of antiviral effect. C-terminal viperin mutants still localized to the lipid droplet interface and replication complexes but did not interact with NS5A proteins as determined by FRET analysis. In conclusion we propose that viperin interacts with NS5A and the host factor VAP-A to limit HCV replication at the replication complex. This highlights the complexity of host control of viral replication by interferon stimulated gene expression

Viperin is induced following dengue virus type-2 (DENV-2) infection and has anti-viral actions requiring the C-terminal end of viperin

The host protein viperin is an interferon stimulated gene (ISG) that is up-regulated during a number of viral infections. In this study we have shown that dengue virus type-2 (DENV-2) infection significantly induced viperin, co-incident with production of viral RNA and via a mechanism requiring retinoic acid-inducible gene I (RIG-I). Viperin did not inhibit DENV-2 entry but DENV-2 RNA and infectious virus release was inhibited in viperin expressing cells. Conversely, DENV-2 replicated to higher tires earlier in viperin shRNA expressing cells. The anti-DENV effect of viperin was mediated by residues within the C-terminal 17 amino acids of viperin and did not require the N-terminal residues, including the helix domain, leucine zipper and S-adenosylmethionine (SAM) motifs known to be involved in viperin intracellular membrane association. Viperin showed co-localisation with lipid droplet markers, and was co-localised and interacted with DENV-2 capsid (CA), NS3 and viral RNA. The ability of viperin to interact with DENV-2 NS3 was associated with its anti-viral activity, while co-localisation of viperin with lipid droplets was not. Thus, DENV-2 infection induces viperin which has anti-viral properties residing in the C-terminal region of the protein that act to restrict early DENV-2 RNA production/accumulation, potentially via interaction of viperin with DENV-2 NS3 and replication complexes. These anti-DENV-2 actions of viperin show both contrasts and similarities with other described anti-viral mechanisms of viperin action and highlight the diverse nature of this unique anti-viral host protein.Karla J. Helbig, Jillian M. Carr, Julie K. Calvert, Satiya Wati, Jennifer N. Clarke, Nicholas S. Eyre, Sumudu K. Narayana, Guillaume N. Fiches, Erin M. McCartney, Michael R. Bear

Inhibition of Tip60 Reduces Lytic and Latent Gene Expression of Kaposi’s Sarcoma-Associated Herpes Virus (KSHV) and Proliferation of KSHV-Infected Tumor Cells

Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic virus responsible for the development of Kaposi’s sarcoma, primary effusion lymphoma (PEL), and Multicentric Castleman’s disease in immunocompromised individuals. Despite the burden of these diseases there are few treatment options for afflicted individuals, due in part to our limited understanding of virus-host interactions. Tip60, a histone aceytltransferase (HAT) has been previously shown to interact with both the KSHV latency associated nuclear antigen protein (LANA), which is the main factor in maintaining the viral latent state, and ORF36, a viral kinase expressed in the lytic phase. We further investigated Tip60-virus interaction to ascertain Tip60’s role in the viral life cycle and its potential as a target for future therapeutics. Through modulation of Tip60 expression in HEK293T cells harboring a plasmid containing the KSHV viral episome, Bac36, we found that Tip60 is vital for both lytic replication as well as efficient expression of latent genes. Interestingly, Tip60 small molecule inhibitors, MG149 and NU9056, similarly inhibited latent and lytic genes, and reduced virion production in wild-type KSHV+/EBV- PEL, BCBL-1 cells. Long-term treatment with these Tip60 inhibitors selectively decreased the viability of KSHV-infected B lymphoma cells compared to uninfected cells. From this study, we conclude that Tip60 is important for KSHV infection and its associated cancer development, and Tip60 is therefore a potential target for future antiviral and anticancer therapeutics

Diversified Application of Barcoded PLATO (PLATO-BC) Platform for Identification of Protein Interactions

Proteins usually associate with other molecules physically to execute their functions. Identifying these interactions is important for the functional analysis of proteins. Previously, we reported the parallel analysis of translated ORFs (PLATO) to couple ribosome display of full-length ORFs with affinity enrichment of mRNA/protein/ribosome complexes for the “bait” molecules, followed by the deep sequencing analysis of mRNA. However, the sample processing, from extraction of precipitated mRNA to generation of DNA libraries, includes numerous steps, which is tedious and may cause the loss of materials. Barcoded PLATO (PLATO-BC), an improved platform was further developed to test its application for protein interaction discovery. In this report, we tested the antisera-antigen interaction using serum samples from patients with inclusion body myositis (IBM). Tripartite motif containing 21 (TRIM21) was identified as a potentially new IBM autoantigen. We also expanded the application of PLATO-BC to identify protein interactions for JQ1, single ubiquitin peptide, and NS5 protein of Zika virus. From PLATO-BC analyses, we identified new protein interactions for these “bait” molecules. We demonstrate that Ewing sarcoma breakpoint region 1 (EWSR1) binds to JQ1 and their interactions may interrupt the EWSR1 association with acetylated histone H4. RIO kinase 3 (RIOK3), a newly identified ubiquitin-binding protein, is preferentially associated with K63-ubiquitin chain. We also find that Zika NS5 protein interacts with two previously unreported host proteins, par-3 family cell polarity regulator (PARD3) and chromosome 19 open reading frame 53 (C19orf53), whose attenuated expression benefits the replication of Zika virus. These results further demonstrate that PLATO-BC is capable of identifying novel protein interactions for various types of “bait” molecules. Keywords: Barcoded PLATO, Protein interaction, Ubiquitin-binding protein, Bromodomain inhibitor JQ1, Zika viru

Data_Sheet_1_Inhibition of Tip60 Reduces Lytic and Latent Gene Expression of Kaposi’s Sarcoma-Associated Herpes Virus (KSHV) and Proliferation of KSHV-Infected Tumor Cells.docx

<p>Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic virus responsible for the development of Kaposi’s sarcoma, primary effusion lymphoma (PEL), and Multicentric Castleman’s disease in immunocompromised individuals. Despite the burden of these diseases there are few treatment options for afflicted individuals, due in part to our limited understanding of virus-host interactions. Tip60, a histone aceytltransferase (HAT) has been previously shown to interact with both the KSHV latency associated nuclear antigen protein (LANA), which is the main factor in maintaining the viral latent state, and ORF36, a viral kinase expressed in the lytic phase. We further investigated Tip60-virus interaction to ascertain Tip60’s role in the viral life cycle and its potential as a target for future therapeutics. Through modulation of Tip60 expression in HEK293T cells harboring a plasmid containing the KSHV viral episome, Bac36, we found that Tip60 is vital for both lytic replication as well as efficient expression of latent genes. Interestingly, Tip60 small molecule inhibitors, MG149 and NU9056, similarly inhibited latent and lytic genes, and reduced virion production in wild-type KSHV⁺/EBV^- PEL, BCBL-1 cells. Long-term treatment with these Tip60 inhibitors selectively decreased the viability of KSHV-infected B lymphoma cells compared to uninfected cells. From this study, we conclude that Tip60 is important for KSHV infection and its associated cancer development, and Tip60 is therefore a potential target for future antiviral and anticancer therapeutics.</p

Data_Sheet_2_Inhibition of Tip60 Reduces Lytic and Latent Gene Expression of Kaposi’s Sarcoma-Associated Herpes Virus (KSHV) and Proliferation of KSHV-Infected Tumor Cells.docx

<p>Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic virus responsible for the development of Kaposi’s sarcoma, primary effusion lymphoma (PEL), and Multicentric Castleman’s disease in immunocompromised individuals. Despite the burden of these diseases there are few treatment options for afflicted individuals, due in part to our limited understanding of virus-host interactions. Tip60, a histone aceytltransferase (HAT) has been previously shown to interact with both the KSHV latency associated nuclear antigen protein (LANA), which is the main factor in maintaining the viral latent state, and ORF36, a viral kinase expressed in the lytic phase. We further investigated Tip60-virus interaction to ascertain Tip60’s role in the viral life cycle and its potential as a target for future therapeutics. Through modulation of Tip60 expression in HEK293T cells harboring a plasmid containing the KSHV viral episome, Bac36, we found that Tip60 is vital for both lytic replication as well as efficient expression of latent genes. Interestingly, Tip60 small molecule inhibitors, MG149 and NU9056, similarly inhibited latent and lytic genes, and reduced virion production in wild-type KSHV⁺/EBV^- PEL, BCBL-1 cells. Long-term treatment with these Tip60 inhibitors selectively decreased the viability of KSHV-infected B lymphoma cells compared to uninfected cells. From this study, we conclude that Tip60 is important for KSHV infection and its associated cancer development, and Tip60 is therefore a potential target for future antiviral and anticancer therapeutics.</p

Viperin protein is induced in DENV-2 infected cells.

<p><b>A.</b> Primary MDM were left uninfected, treated with 500 U/ml IFN-α or DENV-2 infected. At 48 h pi cells were lysed and viperin protein analysed by western blot. Blots were re-probed for β-actin and images visualised by chemiluminesence. Images were quantitated using Carestream Molecular Imaging Software and viperin signal normalised against β-actin. <b>B.</b> Primary MDM were DENV-2 (i) or mock (ii) infected and at 24 h pi were fixed and immunostained for viperin and DENV, with detection of stained complexes with anti-rabbit 647 (red) and anti-mouse 488 (green), respectively. Nuclei were stained with Hoechst (blue) and images collected by confocal microscopy. <b>C.</b> Immunolabeling for viperin was quantitated in cells from mock-infected MDM and compared with antigen negative bystander and DENV-2 antigen positive cells of the DENV-2 infected MDM cultures. Values represent average ± SEM. (n = 111 mock; 27 DENV-antigen positive; 136 DENV-antigen negative bystander cells). * = significantly different, p<0.05, Students unpaired t-test. Results of a single experiment are shown which was replicated.</p