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Dissertation
Regulation of HIV-1 LTR driven gene expression within a chromatin-based microenvironment
Doctor of Philosophy (Ph.D.), Drexel University
Apr 2012
DOI:
https://doi.org/10.17918/00007559
Abstract
Human immunodeficiency virus type 1 (HIV-1) gene expression is driven by the long terminal repeat (LTR), which contains binding sites that interact with multiple host and viral factors. Selective pressures within the host as well as the low efficiency of reverse transcriptase lead to genetic alterations within the viral genome resulting in viral quasispecies that can be differentially regulated and can potentially form niches within specific cell types and tissues. Previous studies identified a single nucleotide polymorphism (SNP) within C/EBP site I (3T, C-to-T change at position 3 of the site) that correlated with HIV-1-associated dementia. In addition, our current patient cohort shows a SNP within Sp site III (5T, C-to-T change at position 5 of the site) that occurs as frequently as the consensus subtype B sequence. Stably transfected cell lines were developed using bone marrow progenitor, T, and monocytic cell lines (TF-1, Jurkat, and U-937, respectively) to explore the LTR phenotype associated with these genotypic changes from an integrated microenvironment. The LAI LTRs were coupled to the green fluorescent protein (GFP), and polyclonal HIV-1 LTR-GFP stable cell lines were developed. In the stably transfected TF-1 and U-937 cells the 3T/5T-containing LTR was associated with a lower expressing phenotype, that can be induced to high levels of gene expression, following stimulation and/or in the presence of Tat. To examine the mechanism of basal and stimulated LTR driven gene expression, as well as epigenetic modifications that may control it, clones were derived from each population of cells. The clones were examined with respect to basal transcription, cytokine treatment, Tat transactivation, and epigenetic controls. Results suggest that non-expressing clones within monocytic cell lines cannot be induced to express under all conditions. However, the nonexpressing LAI 3T and LAI 5T Jurkat clone genotypes could be induced into expression with both proinflammatory cytokine treatment as well as with epigenetic modification. Results demonstrate that epigenetic modifications to viral and host DNA, cellular phenotype, and genetic variation may determine the overall level of LTR activity and potential to be activated from a quiescent state.
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Details
- Title
- Regulation of HIV-1 LTR driven gene expression within a chromatin-based microenvironment
- Creators
- Sonia Shah
- Contributors
- Brian Wigdahl (Advisor) - Drexel University, Drexel University (1970-)
- Awarding Institution
- Drexel University
- Degree Awarded
- Doctor of Philosophy (Ph.D.)
- Publisher
- Drexel University; Philadelphia, Pennsylvania
- Number of pages
- xxix, 306 pages
- Resource Type
- Dissertation
- Language
- English
- Academic Unit
- Microbiology and Immunology; College of Medicine; Drexel University
- Other Identifier
- 991021888955904721