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Pichia --- Pichia --- DNA. --- DNA --- Genetic polymorphism --- Genetic polymorphism

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Cattle --- Milk production --- Genetic polymorphism --- Selection

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Repression of viral expression is a major strategy developed by animal oncoviruses like BLV (Bovine Leukemia Virus) and HTLV (Human T-cell Leukemia Virus) to escape from the host immune response. Induction of BLV transcription involves the interaction of the viral-encoded Tax protein with the CREB/ATF factors, the resulting complex being able to interact with three 21-bp Tax-responsive elements (TxRE) located in the 5'LTR. These TxREs contain cyclic-AMP responsive elements (CRE) but, remarkably, the "TGACGTCA" consensus is never strictly conserved in any viral strain (AGACGTCA, TGACGGCA, TGACCTCA). To assess the role of these sub-optimal CREs, we introduced a perfect consensus sequence within the TxRE. The trans-activation of a luciferase-based reporter by Tax was not affected in transient transfection assays. Still, in the absence of Tax, basal promoter activity of the mutated LTR was remarkably increased and this enhancement was associated with an increase in the binding efficienc y of the CREB/ATF proteins. In contrast, mutation of other regulatory elements within the LTR (E-box, NFkB, GRE or IRF) did not induce a similar alteration of the basal transcription levels. To evaluate the biological relevance of these observations made in vitro, the mutations were introduced in an infectious BLV molecular clone. After injection into sheep, it appeared that all the recombinants were infectious in vivo and all of them, but one, propagated at wild-type levels. The sole exception was the CRE mutant ; the proviral loads being drastically reduced in the sheep infected with this type of virus. In order to further characterize the mechanisms implicated in activation of viral expression, we aimed to unravel the role of deacetylation. Our approach is based on the use of two highly specific deacetylase inhibitors, trichostatin A (TSA) and trapoxin (TPX). Our results demonstrate that treatment with TSA efficiently enhanced LTR-directed gene expression of integrated reporter cons tructs in heterologous D17 cell lines. To further examine the biological relevance of these observations made in vitro, we analyzed ex vivo isolated peripheral blood mononuclear cells (PBMCs) from BLV-infected sheep. TSA induced a drastic increase in viral expression at levels comparable to PMA + ionomycin, the most efficient activators of BLV expression known to date. Inhibition of deacetylation after addition of TSA or TPX also significantly increased viral expression in PBMCs from cattle, the natural hosts for BLV.

Bovine leukosis --- Bovine leukosis --- Genetic polymorphism --- Genetic polymorphism --- mutation. --- mutation --- Pathogenicity --- Pathogenicity --- gene expression --- gene expression

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DNA. --- DNA --- Bromeliaceae --- Bromeliaceae --- Bacteria --- Bacteria --- PCR --- PCR --- identification. --- identification --- Genetic polymorphism --- Genetic polymorphism --- Billbergia magnifica

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Phaseolus --- Phaseolus --- Genetic polymorphism --- Genetic polymorphism --- Phylogeny --- Phylogeny --- PCR --- PCR --- comminution --- comminution --- DNA. --- DNA