Identification of Cellular Genes Targeted by KSHV-Encoded MicroRNAs

MicroRNAs (miRNAs) are 19 to 23 nucleotide–long RNAs that post-transcriptionally regulate gene expression. Human cells express several hundred miRNAs which regulate important biological pathways such as development, proliferation, and apoptosis. Recently, 12 miRNA genes have been identified within the genome of Kaposi sarcoma–associated herpesvirus; however, their functions are still unknown. To identify host cellular genes that may be targeted by these novel viral regulators, we performed gene expression profiling in cells stably expressing KSHV-encoded miRNAs. Data analysis revealed a set of 81 genes whose expression was significantly changed in the presence of miRNAs. While the majority of changes were below 2-fold, eight genes were down-regulated between 4- and 20-fold. We confirmed miRNA-dependent regulation for three of these genes and found that protein levels of thrombospondin 1 (THBS1) were decreased >10-fold. THBS1 has previously been reported to be down-regulated in Kaposi sarcoma lesions and has known activity as a strong tumor suppressor and anti-angiogenic factor, exerting its anti-angiogenic effect in part by activating the latent form of TGF-β. We show that reduced THBS1 expression in the presence of viral miRNAs translates into decreased TGF-β activity. These data suggest that KSHV-encoded miRNAs may contribute directly to pathogenesis by down-regulation of THBS1, a major regulator of cell adhesion, migration, and angiogenesis

Upregulation of xCT by KSHV-Encoded microRNAs Facilitates KSHV Dissemination and Persistence in an Environment of Oxidative Stress

Upregulation of xCT, the inducible subunit of a membrane-bound amino acid transporter, replenishes intracellular glutathione stores to maintain cell viability in an environment of oxidative stress. xCT also serves as a fusion-entry receptor for the Kaposi's sarcoma-associated herpesvirus (KSHV), the causative agent of Kaposi's sarcoma (KS). Ongoing KSHV replication and infection of new cell targets is important for KS progression, but whether xCT regulation within the tumor microenvironment plays a role in KS pathogenesis has not been determined. Using gene transfer and whole virus infection experiments, we found that KSHV-encoded microRNAs (KSHV miRNAs) upregulate xCT expression by macrophages and endothelial cells, largely through miR-K12-11 suppression of BACH-1—a negative regulator of transcription recognizing antioxidant response elements within gene promoters. Correlative functional studies reveal that upregulation of xCT by KSHV miRNAs increases cell permissiveness for KSHV infection and protects infected cells from death induced by reactive nitrogen species (RNS). Interestingly, KSHV miRNAs simultaneously upregulate macrophage secretion of RNS, and biochemical inhibition of RNS secretion by macrophages significantly reduces their permissiveness for KSHV infection. The clinical relevance of these findings is supported by our demonstration of increased xCT expression within more advanced human KS tumors containing a larger number of KSHV-infected cells. Collectively, these data support a role for KSHV itself in promoting de novo KSHV infection and the survival of KSHV-infected, RNS-secreting cells in the tumor microenvironment through the induction of xCT

Regulation and Autoregulation of the Promoter for the Latency-associated Nuclear Antigen of Kaposi's Sarcoma-associated Herpesvirus

Kaposi's sarcoma-associated herpesvirus (KSHV) or human herpesvirus 8 has been established as the etiological agent of Kaposi's sarcoma and certain AIDS-associated lymphomas. KSHV establishes latent infection in these tumors, invariably expressing high levels of the viral latency-associated nuclear antigen (LANA) protein. LANA is necessary and sufficient to maintain the KSHV episome. It also modulates viral and cellular transcription and has been implicated directly in oncogenesis because of its ability to bind to the p53 and pRb tumor suppressor proteins. Previously, we identified the LANA promoter (LANAp) and showed that it was positively regulated by LANA itself. Here, we present a detailed mutational analysis and define cis-acting elements and trans-acting factors for the core LANAp. We found that a downstream promoter element, TATA box, and GC box/Sp1 site at -29 are all individually required for activity. This architecture places LANAp into the small and unusual group of eukaryotic promoters that contain both the downstream promoter element and TATA element but lack a defined initiation site. Furthermore, we demonstrate that LANA regulates its own promoter via its C-terminal domain and does bind to a defined site within the core promoter

PDGFRA defines the mesenchymal stem cell Kaposi's sarcoma progenitors by enabling KSHV oncogenesis in an angiogenic environment

Kaposi’s sarcoma (KS) is an AIDS-defining cancer caused by the KS-associated herpesvirus (KSHV). Unanswered questions regarding KS are its cellular ontology and the conditions conducive to viral oncogenesis. We identify PDGFRA(+)/SCA-1(+) bone marrow-derived mesenchymal stem cells (Pα(+)S MSCs) as KS spindle-cell progenitors and found that pro-angiogenic environmental conditions typical of KS are critical for KSHV sarcomagenesis. This is because growth in KS-like conditions generates a de-repressed KSHV epigenome allowing oncogenic KSHV gene expression in infected Pα(+)S MSCs. Furthermore, these growth conditions allow KSHV-infected Pα(+)S MSCs to overcome KSHV-driven oncogene-induced senescence and cell cycle arrest via a PDGFRA-signaling mechanism; thus identifying PDGFRA not only as a phenotypic determinant for KS-progenitors but also as a critical enabler for viral oncogenesis.Fil: Naipauer, Julian. Miami University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados UnidosFil: Rosario, Santas. Miami University; Estados Unidos. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados UnidosFil: Gupta, Sachin. Miami University; Estados Unidos. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados UnidosFil: Premer, Courtney. Miami University; Estados UnidosFil: Méndez Solís, Omayra. Miami University; Estados Unidos. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados UnidosFil: Schlesinger, Mariana. Miami University; Estados Unidos. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Ponzinibbio, Maria Virginia. Miami University; Estados Unidos. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Jain, Vaibhav. University of Florida; Estados UnidosFil: Gay, Lauren. University of Florida; Estados UnidosFil: Renne, Rolf. University of Florida; Estados UnidosFil: Chan, Ho Lam. Miami University; Estados UnidosFil: Morey, Lluis. Miami University; Estados UnidosFil: Salyakina, Daria. Miami University; Estados Unidos. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados UnidosFil: Abba, Martín Carlos. Miami University; Estados Unidos. Universidad Nacional de La Plata. Facultad de Ciencias Médicas. Centro de Investigaciones Inmunológicas Básicas y Aplicadas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Williams, Sion. Miami University; Estados UnidosFil: Hare, Joshua M.. Miami University; Estados UnidosFil: Goldschmidt Clermont, Pascal. Miami University; Estados UnidosFil: Mesri, Enrique Alfredo. Sylvester Comprehensive Cancer Center and Miami Center for AIDS Research; Estados Unidos. Miami University; Estados Unido

Fucino palaeo-lake: Towards the Palaeoenvironmental history of the last 430 Ka

© 2010 AIQUA - Associazione Italiana per lo Studio del Quaternario e EMMEVI - Servizio Congressi SPA. The sedimentary succession deposited in Fucino palaeo-lake potentially records the environmental history of the Central Mediterranean Region continuously since the early Pleistocene and up to recent historical times. Fucino palaeo-lake sediments are interbedded with numerous volcanic ash layers which allow the reconstruction a robust and independent chronological framework of past environment changes. This framework is a fundamental tool to synchronise different archives at a regional and extra-regional scale and to better understand the spatio-temporal climate variability in the Quaternary at the orbital and millennial-scales. Here we present new preliminary data for the last five glacial to interglacial cycles

Extending the tephra and palaeoenvironmental record of the Central Mediterranean back to 430 ka: A new core from Fucino Basin, central Italy

Here we present the first tephrostratigraphic, palaeomagnetic, and multiproxy data from a new ∼98 m-deep sediment core retrieved from the Fucino Basin, central Italy, spanning the last ∼430 kyr. Palaeoenvironmental proxy data (Ca-XRF, gamma ray and magnetic susceptibility) show a cyclical variability related to interglacial-glacial cycles since the Marine Isotope Stage (MIS) 12-MIS 11 transition. More than 130 tephra layers are visible to the naked eye, 11 of which were analysed (glass-WDS) and successfully correlated to known eruptions and/or other equivalent tephra. In addition to tephra already recognised in the previously investigated cores spanning the last 190 kyr, we identified for the first time tephra from the eruptions of: Tufo Giallo di Sacrofano, Sabatini (288.0 ± 2.0 ka); Villa Senni, Colli Albani (367.5 ± 1.6 ka); Pozzolane Nere and its precursor, Colli Albani (405.0 ± 2.0 ka, and 407.1 ± 4.2 ka, respectively) and Castel Broco, Vulsini (419–490 ka). The latter occurs at the bottom of the core and has been 40Ar/39Ar dated at 424.3 ± 3.2 ka, thus providing a robust chronological constrain for both the eruption itself and the base of the investigated succession. Direct 40Ar/39Ar dating and tephra geochemical fingerprinting provide a preliminary radioisotopic-based chronological framework for the MIS 11-MIS 7 interval, which represent a foundation for the forthcoming multiproxy studies and for investigating the remaining ∼110 tephra layers that are recorded within this interval. Such future developments will contribute towards an improved MIS 11-MIS 7 Mediterranean tephrostratigraphy, which is still poorly explored and exploited