| The Human T-cell leukemia virus type 1 (HTLV-1) and type 2 (HTLV-2) are two pathogenic human retroviruses that are closely related at the amino acid sequence and genome organization, but highly distinct in their pathogenesis. In vitro, they both have the capacity to transform human primary T cells. On the other hand, HTLV-1 is the causative agent for adult T-cell leukemia lymphoma and HTLV-1 associated myelopathy, whereas HTLV-2 is less pathogenic and has been reported to be endemic in intravenous drug users. In this dissertation, we report molecular studies regarding the regulation of HTLV replication and its impact on viral persistence in vivo. In Chapter 2, we generate a novel HTLV-1 clone (H1IT) in which the two regulatory proteins, Tax and Rex, have been separated in an attempt to provide a better reagent to study mutants of these proteins in the context of the provirus and analyze their contribution to HTLV-mediated transformation. Transient transfection of H1IT shows that it expresses both functional Tax and Rex and is able to produce p19Gag. In short and long-term coculture assays, data indicate that H1IT is replication competent and is capable of cellular transformation of primary human T-cells. On the other hand, H1IT was not able to persist in vivo, emphasizing the importance of temporal and quantitative regulation of specific RNA to viral replication in vitro and in vivo.;In Chapter 3, we report that both HTLV-1 and HTLV-2 have evolved accessory genes whose products are able to restrict viral replication at a post-transcriptional level. The HTLV-1 p30II and the related HTLV-2 p28II proteins act as negative regulators of both Tax and Rex by binding to and retaining tax/rex mRNA in the nucleus, thereby inhibiting virion production. Reduction of viral replication in a cell carrying the provirus may allow escape from immune recognition in an infected individual.;In Chapter 4, we follow up on data in Chapter 3 to show that p28 II is recruited to the viral promoter in a Tax-dependent manner. After recruitment to the promoter, p28II or p30II then travels with the transcription elongation machinery until its target mRNA is synthesized. The result of experiments artificially directing these proteins to the promoter indicated that p28II, unlike HTLV-1 p30II displays no transcriptional activity. Furthermore, the tethering of p28II directly to tax/rex mRNA resulted in repression of Tax function.;Since data in Chapters 3 and 4 are consistent with a critical role of these accessory proteins in viral persistence in vivo, in Chapter 5, we used an animal model of HTLV-1 and HTLV-2 infection to study the specific contribution of p28II on HTLV-2 survival in rabbits. In this study, all wtHTLV-2 infected rabbits showed persistent infection, whereas those infected with HTLV-2Deltap28 were able to eliminate the virus as early as 2 weeks, indicating that p28II is critical for early viral infectivity, spread and/or persistence in infected rabbits. Collectively, data presented within this thesis support the conclusion that the regulation of HTLV gene expression a complicated but a tightly controlled process. |
