Mathieu Boxus

The HTLV-1 Tax interactome.

The Tax1 oncoprotein encoded by Human T-lymphotropic virus type I is a major determinant of viral persistence and pathogenesis. Tax1 affects a wide variety of cellular signalling pathways leading to transcriptional activation, proliferation and ultimately transformation. To carry out these functions, Tax1 interacts with and modulates activity of a number of cellular proteins. In this review, we summarize the present knowledge of the Tax1 interactome and propose a rationale for the broad range of cellular proteins identified so far.

Mechanisms of leukemogenesis induced by bovine leukemia virus: prospects for novel anti-retroviral therapies in human

In 1871, the observation of yellowish nodules in the enlarged spleen of a cow was considered to be the first reported case of bovine leukemia. The etiological agent of this lymphoproliferative disease, bovine leukemia virus (BLV), belongs to the deltaretrovirus genus which also includes the related human T-lymphotropic virus type 1 (HTLV-1). This review summarizes current knowledge of this viral system, which is important as a model for leukemogenesis. Recently, the BLV model has also cast light onto novel prospects for therapies of HTLV induced diseases, for which no satisfactory treatment exists so far.

Mechanisms of HTLV-1 persistence and transformation.

Adult T-cell leukaemia (ATL) is caused by the human T-cell lymphotropic virus type 1 (HTLV-1). HTLV-1 has elaborated strategies to persist and replicate in the presence of a strong immune response. In this review, we summarise these mechanisms and their contribution to T-cell transformation and ATL development.

Preventive and Therapeutic Strategies for Bovine Leukemia Virus: Lessons for HTLV

Bovine leukemia virus (BLV) is a retrovirus closely related to the human T-lymphotropic virus type 1 (HTLV-1). BLV is a major animal health problem worldwide causing important economic losses. A series of attempts were developed to reduce prevalence, chiefly by eradication of infected cattle, segregation of BLV-free animals and vaccination. Although having been instrumental in regions such as the EU, these strategies were unsuccessful elsewhere mainly due to economic costs, management restrictions and lack of an efficient vaccine. This review, which summarizes the different attempts previously developed to decrease seroprevalence of BLV, may be informative for management of HTLV-1 infection. We also propose a new approach based on competitive infection with virus deletants aiming at reducing proviral loads.

Host-Pathogen Interactome Mapping for HTLV-1 and -2 Retroviruses

BackgroundHuman T-cell leukemia virus type 1 (HTLV-1) and type 2 both target T lymphocytes, yet induce radically different phenotypic outcomes. HTLV-1 is a causative agent of Adult T-cell leukemia (ATL), whereas HTLV-2, highly similar to HTLV-1, causes no known overt disease. HTLV gene products are engaged in a dynamic struggle of activating and antagonistic interactions with host cells. Investigations focused on one or a few genes have identified several human factors interacting with HTLV viral proteins. Most of the available interaction data concern the highly investigated HTLV-1 Tax protein. Identifying shared and distinct host-pathogen protein interaction profiles for these two viruses would enlighten how they exploit distinctive or common strategies to subvert cellular pathways toward disease progression.ResultsWe employ a scalable methodology for the systematic mapping and comparison of pathogen-host protein interactions that includes stringent yeast two-hybrid screening and systematic retest, as well as two independent validations through an additional protein interaction detection method and a functional transactivation assay. The final data set contained 166 interactions between 10 viral proteins and 122 human proteins. Among the 166 interactions identified, 87 and 79 involved HTLV-1 and HTLV-2 -encoded proteins, respectively. Targets for HTLV-1 and HTLV-2 proteins implicate a diverse set of cellular processes including the ubiquitin-proteasome system, the apoptosis, different cancer pathways and the Notch signaling pathway.ConclusionsThis study constitutes a first pass, with homogeneous data, at comparative analysis of host targets for HTLV-1 and -2 retroviruses, complements currently existing data for formulation of systems biology models of retroviral induced diseases and presents new insights on biological pathways involved in retroviral infection.

Bioreactor mixing efficiency modulates the activity of a prpoS::GFP reporter gene in E. coli

BackgroundExtensive studies have shown that up-scaling of bioprocesses has a significant impact on the physiology of the microorganisms. Among the factors associated with the fluid dynamics of the bioreactor, concentration gradients induced by loss of the global mixing efficiency associated with the increasing scale is the main phenomena leading to strong physiological modifications at the level of the microbial population. These changes are not fully understood since they involve complex physiological mechanisms. In this work, we intend to investigate, at the single cell level, the expression of the rpoS gene associated with the stress response of E. coli. The cultures of the reporter strain have been performed in a small scale reactor as well as in a series of scaled-down bioreactors able to induce extracellular perturbations with increasing level of magnitude.ResultsThe rpoS level has been monitored by the aim of a transcriptional reporter gene based on the synthesis of the green fluorescent protein (GFP). It has been observed that the level of GFP increases during the transition from batch to fed-batch phase. After this initial increase, the GFP content of the cell drops, primarily due to the dilution by cell division. However, a significant drop of the GFP content has been observed if using a partitioned bioreactor, for which the mixing conditions are very bad, leading to the exposure of the cells to cyclic and stochastic extracellular fluctuations. If considering the flow cytometric profile of the cell to cell GFP content, this drop has to be attributed to the appearance of segregation at the level of the GFP content among the microbial population.ConclusionThe generation of extracellular perturbations (in the present case, at the level of the sugar concentration and the dissolved oxygen level) has led to a drop at the level of the rpoS expression level. This drop has to be attributed to a segregation phenomenon in microbial population, with a major sub-population exhibiting a low expression level and a minor sub-population keeping its initial elevated expression level. The intensity of the segregation, as well as its time of appearance during the culture can be related to the bioreactor mixing efficiency.

Valproate synergizes with purine nucleoside analogues to induce apoptosis of B-chronic lymphocytic leukaemia cells

Resistance to chemotherapy and drug toxicity are two major concerns of chronic lymphocytic leukaemia (B‐CLL) treatment by purine nucleoside analogues (PNA, i.e. fludarabine and cladribine). We hypothesized that targeting epigenetic changes might address these issues and evaluated the effect of the histone deacetylase inhibitor valproate (VPA) at a clinically relevant concentration. VPA acted in a highly synergistic/additive manner with fludarabine and cladribine to induce apoptosis of B‐CLL cells. Importantly, VPA also restored sensitivity to fludarabine in B cells from poor prognosis CLL patients who became resistant to chemotherapy. Mechanism of apoptosis induced by VPA alone or combined with fludarabine or to cladribine was caspase‐dependent and involved the extrinsic pathway. VPA, but neither fludarabine nor cladribine, enhanced the production of reactive oxygen species (ROS) and inhibition of ROS with N‐acetylcysteine decreases apoptosis of CLL cells. VPA stimulates hyperphosphorylation of p42/p44 ERK, cytochrome c release and overexpression of Bax and Fas. Together, our data indicate that VPA may ameliorate the outcome of PNA‐based therapeutic protocols and provide a potential alternative treatment in both the relapsed and front‐line resistant patients and in patients with high risk features.

Cell dynamics and immune response to BLV infection: a unifying model

Bovine Leukemia virus (BLV) is the natural etiological agent of a lymphoproliferative disease in cattle. BLV can also be transmitted experimentally to a related ruminant species, sheep, in which the pathogenesis is more acute. Although both susceptible species develop a strong anti-viral immune response, the virus persists indefinitely throughout life, apparently at a transcriptionally silent stage, at least in a proportion of infected cells. Soon after infection, these humoral and cytotoxic activities very efficiently abolish the viral replicative cycle, permitting only mitotic expansion of provirus-carrying cells. Short term cultures of these infected cells initially indicated that viral expression protects against spontaneous apoptosis, suggesting that leukemia is a process of accumulation of long-lived cells. This conclusion was recently reconsidered following in vivo dynamic studies based on perfusions of nucleoside (bromodeoxyuridine) or fluorescent protein markers (CFSE). In sheep, the turnover rate of infected cells is increased, suggesting that a permanent clearance process is exerted by the immune system. Lymphocyte trafficking from and to the secondary lymphoid organs is a key component in the maintenance of cell homeostasis. The net outcome of the immune selective pressure is that only cells in which the virus is transcriptionally silenced survive and accumulate, ultimately leading to lymphocytosis. Activation of viral and/or cellular expression in this silent reservoir with deacetylase inhibitors causes the collapse of the proviral loads. In other words, modulation of viral expression appears to be curative in lymphocytic sheep, an approach that might also be efficient in patients infected with the related Human T-lymphotropic virus type 1. In summary, a dynamic interplay between BLV and the host immune response modulates a complex equilibrium between (i) viral expression driving (or) favoring proliferation and (ii) viral silencing preventing apoptosis. As conclusion, we propose a hypothetical model unifying all these mechanisms.

The HTLV-1 Tax protein inhibits formation of stress granules by interacting with histone deacetylase 6.

Human T cell leukemia virus type-1 (HTLV-1) is the causative agent of a fatal adult T-cell leukemia. Through deregulation of multiple cellular signaling pathways the viral Tax protein has a pivotal role in T-cell transformation. In response to stressful stimuli, cells mount a cellular stress response to limit the damage that environmental forces inflict on DNA or proteins. During stress response, cells postpone the translation of most cellular mRNAs, which are gathered into cytoplasmic mRNA-silencing foci called stress granules (SGs) and allocate their available resources towards the production of dedicated stress-management proteins. Here we demonstrate that Tax controls the formation of SGs and interferes with the cellular stress response pathway. In agreement with previous reports, we observed that Tax relocates from the nucleus to the cytoplasm in response to environmental stress. We found that the presence of Tax in the cytoplasm of stressed cells prevents the formation of SGs and counteracts the shutoff of specific host proteins. Unexpectedly, nuclear localization of Tax promotes spontaneous aggregation of SGs, even in the absence of stress. Mutant analysis revealed that the SG inhibitory capacity of Tax is independent of its transcriptional abilities but relies on its interaction with histone deacetylase 6, a critical component of SGs. Importantly, the stress-protective effect of Tax was also observed in the context of HTLV-1 infected cells, which were shown to be less prone to form SGs and undergo apoptosis under arsenite exposure. These observations identify Tax as the first virally encoded inhibitory component of SGs and unravel a new strategy developed by HTLV-1 to deregulate normal cell processes. We postulate that inhibition of the stress response pathway by Tax would favor cell survival under stressful conditions and may have an important role in HTLV-1-induced cellular transformation.

HDAC5 is required for maintenance of pericentric heterochromatin, and controls cell-cycle progression and survival of human cancer cells

Histone deacetylases (HDACs) form a family of enzymes, which have fundamental roles in the epigenetic regulation of gene expression and contribute to the growth, differentiation, and apoptosis of cancer cells. In this study, we further investigated the biological function of HDAC5 in cancer cells. We found HDAC5 is associated with actively replicating pericentric heterochromatin during late S phase. We demonstrated that specific depletion of HDAC5 by RNA interference resulted in profound changes in the heterochromatin structure and slowed down ongoing replication forks. This defect in heterochromatin maintenance and assembly are sensed by DNA damage checkpoint pathways, which triggered cancer cells to autophagy and apoptosis, and arrested their growth both in vitro and in vivo. Finally, we also demonstrated that HDAC5 depletion led to enhanced sensitivity of DNA to DNA-damaging agents, suggesting that heterochromatin de-condensation induced by histone HDAC5 silencing may enhance the efficacy of cytotoxic agents that act by targeting DNA in vitro. Together, these results highlighted for the first time an unrecognized link between HDAC5 and the maintenance/assembly of heterochromatin structure, and demonstrated that its specific inhibition might contribute to increase the efficacy of DNA alteration-based cancer therapies in clinic.