Thesis Format
Integrated Article
Degree
Doctor of Philosophy
Program
Microbiology and Immunology
Supervisor
Dikeakos, Jimmy D.
Abstract
Human Immunodeficiency Virus Type 1 (HIV-1) expresses viral proteins that mediate disease progression. My thesis focuses on the HIV-1 accessory protein Nef, a viral protein that facilitates host immune evasion and enhances viral infectivity in people with HIV-1 (PWH). These functions are mediated by Nef’s ability to modulate cell surface expression of host human proteins, including Major Histocompatibility Complex Class I (MHC-I), Cluster of Differentiation 4 (CD4), and Serine Incorporator 5 (SERINC5). In my first aim, a bioinformatics pipeline was developed to create a time-scaled phylogeny relating extant primate nef sequences. The nef sequences of all ancestors involved in the phylogeny towards HIV-1 group M were then reconstructed. All Nef ancestors were assessed for SERINC5, CD4, and CD3ζ downregulation to validate the accuracy of the reconstruction and to test how these functions evolved. Functional characterization of all reconstructed Nef ancestors involved in the phylogeny towards HIV-1 group M allowed for estimations of how Nef-mediated SERINC5, CD4, and CD3ζ downregulation changed throughout its evolutionary history, thereby demonstrating this pipeline as a viable strategy to estimate how other viral functions may have evolved. In the second aim, I defined Nef-mediated SERINC5 and CD4 downregulation in a cohort of PWH. I found that two primary Nef proteins drastically differed in their SERINC5 downregulatory capabilities yet retained the ability to downregulate CD4. My studies mapped the Nef region responsible for this functional uncoupling to the highly conserved C-terminal dileucine motif. Identification of this polymorphism provides clues regarding the biochemical nature of the Nef:SERINC5 interaction and the pathways Nef hijacks to antagonize SERINC5. In the third aim, I explored whether Nef-mediated MHC-I downregulation contributes to the size of the replication-competent latent viral reservoir (RC-LVR) in PWH on long-term antiretroviral therapy (ART). Nef-dependent MHC-I downregulation was significantly inversely correlated with the rate of change of the RC-LVR, suggesting that efficient evasion of the cytotoxic T cell (CTL) response contributes to the maintenance of the RC-LVR in PWH. Overall, these studies provide a framework for understanding how Nef functions evolved and how these functions contribute to pathogenesis and virulence in PWH.
Summary for Lay Audience
Human Immunodeficiency Virus Type 1 (HIV-1) remains a global health burden for human populations. One of the proteins HIV-1 makes is called Nef, which helps HIV-1 evade the body’s immune system, thus making it easier for HIV-1 to persist and infect new cells. To do this, Nef counters several host proteins, in particular Major Histocompatibility Complex Class I (MHC-I), Cluster of Differentiation 4 (CD4), Cluster of Differentiation 3ζ (CD3ζ) and Serine Incorporator 5 (SERINC5). My research generally involves understanding how these Nef functions impact HIV-1 virulence in people with HIV (PWH). First, I developed a computer program to understand the genetic history of Nef from HIV and simian immunodeficiency virus (SIV) and determined the sequences of the Nef ancestors. I then tested the ability of these Nef ancestors to counter CD4, SERINC5 and CD3ζ to understand how these Nef functions evolved through time. Second, I determined the ability of Nef proteins from PWH in Uganda to counter SERINC5, which was largely unknown at the start of my graduate studies. I found that two primary Nef proteins were able to counter CD4, but only one was able to counter SERINC5. By swapping segments of the one Nef protein with the other, I was able to determine the region of Nef that was involved in countering SERINC5. Identifying this specific region involved in countering SERINC5 is important for understanding how Nef interacts with SERINC5, which is poorly understood presently. Finally, I looked at the ability of Nef from several PWH on long-term antiretroviral treatment to target MHC-I, a key function allowing HIV to hide from the immune system, and determine if this function is related to changes in the frequency of latently HIV-1 infected cells during treatment. I found that Nef proteins decreasing cell surface MHC-I levels more efficiently resulted in slower decay of the latent reservoir, and vice versa. Overall, my research helps us understand how Nef evolves over time and how it evades the body’s immune system in PWH, which can help us develop more effective treatments for those with HIV-1 accordingly.
Recommended Citation
Mumby, Mitchell J., "Functional and evolutionary dynamics of HIV-1 Nef: Contributions to viral virulence and immune evasion" (2024). Electronic Thesis and Dissertation Repository. 10531.
https://ir.lib.uwo.ca/etd/10531
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