Showing papers by "Benoit Barbeau published in 2012"

Human T-cell leukemia virus type 1 (HTLV-1) bZIP factor requires cellular transcription factor JunD to upregulate HTLV-1 antisense transcription from the 3' long terminal repeat.

Abstract: Infection with the human T-cell leukemia virus type 1 (HTLV-1) results in a variety of diseases including adult T-cell leukemia (ATL), a fatal malignancy characterized by the uncontrolled proliferation of virally infected CD4(+) T cells. The HTLV-1 basic leucine zipper factor (HBZ) is believed to contribute to development and maintenance of ATL. Unlike the other HTLV-1 genes, the hbz gene is encoded on the complementary strand of the provirus and therefore is not under direct control of the promoter within the 5' long terminal repeat (LTR) of the provirus. This promoter can undergo inactivating genetic or epigenetic changes during the course of ATL that eliminates expression of all viral genes except that of hbz. In contrast, repressive modifications are not known to occur on the hbz promoter located in the 3' LTR, and hbz expression has been consistently detected in all ATL patient samples. Although Sp1 regulates basal transcription from the HBZ promoter, other factors that activate transcription remain undefined. In this study, we used a proviral reporter construct deleted of the 5' LTR to show that HBZ upregulates its own expression through cooperation with JunD. Activation of antisense transcription was apparent in serum-deprived cells in which the level of JunD was elevated, and elimination of JunD expression by gene knockout or shRNA-mediated knockdown abrogated this effect. Activation through HBZ and JunD additionally required Sp1 binding at the hbz promoter. These data favor a model in which JunD is recruited to the promoter through Sp1, where it heterodimerizes with HBZ thereby enhancing its activity. Separately, hbz gene expression led to an increase in JunD abundance, and this effect correlated with emergence of features of transformed cells in immortalized fibroblasts. Overall, our results suggest that JunD represents a novel therapeutic target for the treatment of ATL.

HIV-1 Antisense Transcription Is Preferentially Activated in Primary Monocyte-Derived Cells

Abstract: In this study, an antisense luciferase-expressing human immunodeficiency virus type 1 (HIV-1) molecular clone was used to infect primary cells. We found that antisense transcription activity from the 3' long terminal repeat (LTR) was significantly more abundant in monocyte-derived cells than in activated T lymphocytes. Moreover, by analyzing antisense transcription in infected monocyte-derived dendritic cells (MDDCs), we observed that the majority of HIV-1-infected MDDCs with significant antisense transcription activity did not produce Gag. We also confirmed that the negative-strand-encoded antisense protein (ASP) was expressed in monocyte-derived cells.

A new luciferase-based quantitative assay for the evaluation of human trophoblast fusion.

Abstract: The syncytiotrophoblast is a multinuclear cell layer maintained through fusion events with cytotrophoblasts and plays a key role in the properties of the placenta. Monitoring fusion in this cell layer is important in studies aimed at understanding its function. We herein propose a new fusion assay based on the transactivating potential of the human immunodeficiency virus type 1 (HIV-1) Tat protein on its promoter present in the long terminal repeat (LTR) region. We used 2 BeWo cell populations, one stably transfected with the HIV-1 LTR positioned upstream of the luciferase gene and the other stably transfected with a Tat expression vector. Both stable cell lines were responsive to Tat-mediated LTR transactivation and demonstrated normal fusion and human chorionic gonadotropin (hCG) secretion upon stimulation. When both BeWo cell lines were cocultured, forskolin-mediated induction of fusion led to an increase in luciferase activity, which was sensitive to anti-syncytin 1 and -2 antibodies and syncytin 2 small interfering RNAs (siRNAs). Similar results were obtained in primary trophoblasts. Our results highlight the effectiveness and accuracy of this new quantification assay for trophoblast fusion.