HTLV-1 Integration into Transcriptionally Active Genomic Regions Is Associated with Proviral Expression and with HAM/TSP

Human T-lymphotropic virus type 1 (HTLV-1) causes leukaemia or chronic inflammatory disease in ∼5% of infected hosts. The level of proviral expression of HTLV-1 differs significantly among infected people, even at the same proviral load (proportion of infected mononuclear cells in the circulation). A high level of expression of the HTLV-1 provirus is associated with a high proviral load and a high risk of the inflammatory disease of the central nervous system known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). But the factors that control the rate of HTLV-1 proviral expression remain unknown. Here we show that proviral integration sites of HTLV-1 in vivo are not randomly distributed within the human genome but are associated with transcriptionally active regions. Comparison of proviral integration sites between individuals with high and low levels of proviral expression, and between provirus-expressing and provirus non-expressing cells from within an individual, demonstrated that frequent integration into transcription units was associated with an increased rate of proviral expression. An increased frequency of integration sites in transcription units in individuals with high proviral expression was also associated with the inflammatory disease HAM/TSP. By comparing the distribution of integration sites in human lymphocytes infected in short-term cell culture with those from persistent infection in vivo, we infer the action of two selective forces that shape the distribution of integration sites in vivo: positive selection for cells containing proviral integration sites in transcriptionally active regions of the genome, and negative selection against cells with proviral integration sites within transcription units

Viral Factors Controlling HTLV-1 Proviral Load and Proviral Expression

A high level of provirus expression and a high proviral load during persistent Human T Lymphotropic Virus Type 1 (HTLV-1) infection are associated with an increased risk of the inflammatory disease HTLV-1 associated myelopathy (HAMITSP). However, the factors that control HTLV-1 proviral expression and proviral load in vivo are not known. In this study, we investigate the. influences of epigenetics and the distribution of proviral integration sites on the control of these factors. Using inhibitors of histone deacetylases (HDACi) and methyl-transferases (MTi), we determined that acetylation controls proviral expression in vivo but epigenetic differences between individuals do not cause proviral load and disease-risk variation. • By analysing proviral integration sites derived from individuals persistently infected with HTLV-1, we identified that the distribution' of integration sites is associated with the probability of provirus expression and thG risk of HAMITSP. As an independent verification of this observation, analysis of integration sites found in provirus-expressing and provirus-non-expre~sing infected cells wit~in an individual found that provirus expression is associated with integration into genes. By comparing the distribution of integration sites from HTLV-1-infected individuals with a set of integration sites derived from a co-culture system in vitro, we infer that the distribution of integration sites in persistent infection is subject to in vivo selection. We identified positive selection for cells containing proviral integrations within transcriptionally active regions of the genome but negative selection against cells containing proviral integration sites in transcription units (Le. genes) in vivo. An investigation into alternative markers of transcriptional activity identified a previously uncharacterised transcriptionally active region of the HTLV-1 genome. Preliminary identification and characterization of a novel non-coding anti-sense transcript (AST2) was carried out. In summary, we have demonstrated that the cellular epigenetic status and the distribution of proviral integration sites are associated with provirus ex'pression in vivo. The interplay of these factors with previously defined parameters, such as the efficiency of the CTL response, determines an individual's provirus expression, proviral load and consequent disease risk during persistent infection by HTLV-1.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Epstein-Barr Virus-Induced Resistance to Drugs That Activate the Mitotic Spindle Assembly Checkpoint in Burkitt's Lymphoma Cells

Epstein-Barr virus (EBV) is associated with a number of human cancers, and latent EBV gene expression has been reported to interfere with cell cycle checkpoints and cell death pathways. Here we show that latent EBV can compromise the mitotic spindle assembly checkpoint and rescue Burkitt's lymphoma (BL)-derived cells from caspase-dependent cell death initiated in aberrant mitosis. This leads to unscheduled mitotic progression, resulting in polyploidy and multi- and/or micronucleation. The EBV latent genes responsible for this phenotype are expressed from the P3HR1 strain of virus and several viruses with similar genomic deletions that remove the EBNA2 gene. Although EBNA2 and the latent membrane proteins are not expressed, the EBNA3 proteins are present in these BL cells. Survival of the EBV-positive cells is not consistently associated with EBV lytic gene expression or with the genes that are expressed in EBV latency I BL cells (i.e., EBNA1, EBERs, and BARTs) but correlates with reduced expression of the cellular proapoptotic BH3-only protein Bim. These data suggest that a subset of latent EBV gene products may increase the likelihood of damaged DNA being inherited because of the impaired checkpoint and enhanced survival capacity. This could lead to greater genetic diversity in progeny cells and contribute to tumorigenesis. Furthermore, since it appears that this restricted latent EBV expression interferes with the responses of Burkitt's lymphoma-derived cells to cytotoxic drugs, the results of this study may have important therapeutic implications in the treatment of some BL

In vivo T lymphocyte dynamics in humans and the impact of human T-lymphotropic virus 1 infection

Human T-lymphotropic virus type 1 (HTLV-1) is a persistent CD4(+) T-lymphotropic retrovirus. Most HTLV-1-infected individuals remain asymptomatic, but a proportion develop adult T cell leukemia or inflammatory disease. It is not fully understood how HTLV-1 persists despite a strong immune response or what determines the risk of HTLV-1-associated diseases. Until recently, it has been difficult to quantify lymphocyte kinetics in humans in vivo. Here, we used deuterated glucose labeling to quantify in vivo lymphocyte dynamics in HTLV-1-infected individuals. We then used these results to address four questions. (i) What is the impact of HTLV-1 infection on lymphocyte dynamics? (ii) How does HTLV-1 persist? (iii) What is the extent of HTLV-1 expression in vivo? (iv) What features of lymphocyte kinetics are associated with HTLV-1-associated myelopathy/tropical spastic paraparesis? We found that CD4(+)CD45RO(+) and CD8(+)CD45RO(+) T lymphocyte proliferation was elevated in HTLV-1-infected subjects compared with controls, with an extra 10(12) lymphocytes produced per year in an HTLV-1-infected subject. The in vivo proliferation rate of CD4(+)CD45RO(+) cells also correlated with ex vivo viral expression. Finally, the inflammatory disease HTLV-1-associated myelopathy/tropical spastic paraparesis was associated with significantly increased CD4(+)CD45RO(+) cell proliferation. We suggest that there is persistent viral gene expression in vivo, which is necessary for the maintenance of the proviral load and determines HTLV-1-associated myelopathy/tropical spastic paraparesis risk