Browsing by Subject "Latency"
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Item Open Access Reactive Latency: An Analysis of the Diffusion of Nuclear Latency Between Neighboring States(2019-04-05) McKinney, Katherine E.The threat of a nuclear weapons cascade in the Middle East has perennially plagued US policymakers in their interactions with the region. Accordingly, Herculean efforts have been made to mitigate this threat, and its prevention has been studied extensively. However, Saudi Arabia’s recent interest in pursuing indigenous enrichment capabilities begs a new question: should policymakers be concerned about Iran’s latent status pushing its MENA neighbors to pursue similar capabilities? The threat of reactive latency between neighbors thus demands analysis. Such work is made possible by recent scholarship in the nuclear latency space that aims specifically to support quantitative analysis. In the following article, I contribute to a growing latency- focused literature through an analysis of whether states that have latent neighbors are more likely to become latent themselves. Through three phases of statistical modeling, I analyze the relationships between having a laboratory-scale, pilot-scale, or commercial- scale latent neighbor or neighbors and whether a state itself becomes latent. I find that having a neighbor that has achieved commercial-scale latent capabilities has a positive and nearly statistically significant relationship with whether a state itself becomes latent. This finding could indicate that states may explore nuclear options in response to more modest external proliferation stimuli than is currently believed. Additionally, in many of my models, I find a positive and statistically significant relationship between a state having a nuclear-armed neighbor or neighbors and a state itself becoming latent. This lends further support to the idea that the external proliferation stimuli that beget exploration of and investment in latency may be lower than we had previously thought.Item Open Access Reversal of Epigenetic Silencing by the SMC5/6 Complex Rescues Integrase-Deficient HIV-1 Replication and Prevents the Establishment of a Latent Reservoir.(2022) Darryl irwan, IshakIntegration of HIV-1 DNA is an essential step in the viral life cycle, and how the unintegrated proviral DNA intermediate is transcriptionally silenced has been an ongoing question in the field. Here we show that this transcriptional silencing is epigenetic and demonstrate that the Tax transcription factor encoded by Human T-cell Leukemia Virus 1 (HTLV-1) can reverse the repressive epigenetic modifications on unintegrated, episomal HIV-1 circular DNA. Tax expression in HIV-1 mutants lacking functional integrase, and are normally transcriptionally silenced, causes the recruitment of NF-κB to unintegrated viral DNA promoter, reverses the repressive epigenetic modifications on the chromatinzed viral DNA, and leads to a robust, spreading infection. In addition, using an unbiased screen, we identify the host SMC5/6 complex as essential for epigenetically silencing unintegrated/ pre-integrated HIV-1 DNA. The SMC5/6 complex binds chromatinized HIV-1 DNA and triggers epigenetic silencing by inducing its SUMOylation through NSMCE2, an E3 SUMO ligase. Inhibiting this SUMOylation, by knocking out members of the SMC5/6 complex, mutationally disrupting its E3 ligase function, or by using a SUMOylation inhibitors like TAK-981 prevents this epigenetic silencing and rescues both viral gene transcription from, and replication of, integrase-deficient HIV-1. Finally, we show that inhibiting this initial SUMO-mediated silencing of HIV-1 DNA in integration-competent viruses with a functional integrase by the SMC5/6 complex drastically interferes with the establishment of latent HIV-1 infections in both CD4+ T cell lines and primary human T cells. We thus propose a new model to explain the establishment of the HIV-1 latent reservoir.
Item Open Access Transcriptomic and Metabolic Heterogeneity During Epstein-Barr Virus Latency Establishment(2019) Messinger, Joshua EdwardEpstein-Barr Virus (EBV) is a ubiquitous gamma-herpesvirus in the human population and highly associated with lymphomas of the immune-suppressed. EBV maintains itself within the human host via temporal regulation of viral gene expression before establishing latency in resting memory B cells. Experimentally, we can model these immune-compromised lymphomas via in vitro infection of B cells isolated from peripheral blood. This infection model is biphasic where the initial phase is characterized by hyper-proliferation and expression of the EBV nuclear antigens (EBNAs) in the absence of the latent membrane proteins (LMPs) called latency IIb before transitioning to the NFkB-dependent latency III lymphoblastoid cell line (LCL) stage characterized by full expression of the LMPs in the presence of the EBNAs. This temporal regulation manipulates host cell proliferation rates and metabolic profiles of infected B lymphocytes. Our laboratory and my studies, in particular, are focused on the latency IIb to latency III transition as these states are often observed in patient biopsies of EBV-associated malignancies.
Latent membrane protein 1 (LMP1) expression defines latency III and has been used in lymphoma samples to identify EBV latency III in vivo. While LMP1 expression is lower in latency IIb, the mechanism by which it is repressed is currently unknown. Additionally, while LMP1 expression can be used to distinguish latency IIb from latency III, LMP1 expression varies widely within latency III and a subset of LMP1lo latency III cells express LMP1 at levels similar to latency IIb making these cells hard to distinguish in vitro and in vivo. Lastly, while EBV manipulates host cell metabolism, the viral and host factors important for metabolic rewiring have not been fully elucidated.
Using RT-qPCR, ChIP-qPCR, RNA-sequencing, Western Blotting, flow cytometry, and bioinformatic approaches we have identified c-Myc as a repressor of LMP1 during latency IIb. We have also identified host mRNA biomarkers capable of distinguishing latency IIb from latency III. This discovery was additionally leveraged to develop a multiplex RNA-FISH platform capable of distinguishing latency IIb from latency III. Lastly, we have identified host and viral proteins necessary to rewire host cell metabolism and sustain EBV-mediated B cell proliferation. This work, taken together, adds new understanding to EBV latency establishment, heterogeneity, and host B cell biology to develop new, more targeted, therapeutics for EBV-associated lymphomas in the future.