UBC Theses and Dissertations
Regulation of HIV-1 latency by basal transcription Dahabieh, Matthew Solomon
HIV/AIDS is undoubtedly one of mankind’s most pressing health concerns. Currently, there are ~34 million people infected worldwide, with ~3 million new infections and ~2 million deaths every year. Despite 30 years of research and the development of potent antiretroviral drugs, a cure for HIV-1 remains elusive. This is largely due to viral latency, a phenomenon that makes lifelong HAART therapy essential. While proviral DNA is usually transcribed, the integrated HIV-1 LTR promoter may also exist in a transcriptionally inactive latent state. Current models indicate that latency results primarily from the progressive epigenetic silencing of otherwise active infections. However, the majority of latency models utilize single reporters and selection/culturing to establish latency; therefore, they cannot differentiate between direct and progressive silencing. We hypothesize that direct LTR-silent infections are underappreciated because current models may poorly represent the entire spectrum of HIV-1 latency. In this thesis we aim to characterize a novel double-labeled Red-Green-HIV-1 vector (RGH) to comprehensively study latency. Our results show that, contrary to current dogma, the majority of RGH infections in Jurkat T cells are directly silenced. Moreover, direct silent infections are observed in several cell types and are transcriptionally competent, as known HIV-1 agonists can efficiently reactivate them. We observe that direct silencing occurs at all sites of viral integration and that cellular NFκB levels at the time of infection mediate direct LTR-silencing. Additionally, we aim to characterize the cellular transcription factor RBF-2 with respect to RGH latency and basal transcription. Our results show that RBF-2 binds two conserved sites on the HIV-1 LTR, and that this organization is necessary for mediating proper transcriptional activation. Consequently, this interaction also modulates RGH latency, as RBF-2 mutants displayed higher levels of latency relative to wild type. Collectively, our results shed new light on the previously underappreciated and immeasurable contribution of direct silent infections to HIV-1 latency. Considering these infections make up the majority of total infections in vitro, direct silencing is likely a major component of the latent reservoir in vivo. Fully understanding the entire spectrum of latency, including both direct and progressive mechanisms, will undoubtedly aid HIV-1 eradication strategies.
Item Citations and Data
Attribution-NonCommercial-NoDerivatives 4.0 International