Integration Sites in the Persistence of Latent HIV-1

Author

Amy S. Huang

Date of Award

2022

Document Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

RU Laboratory

Nussenzweig Laboratory

Abstract

Human immunodeficiency virus (HIV-1), the pathogen that causes acquired immune deficiency syndrome (AIDS), remains one of the world’s most pressing health issues. Since the beginning of the HIV/AIDS epidemic, over 32M people have succumbed to AIDS-related illnesses. Despite remarkable advances in HIV-1 biology, neither a vaccine or cure have been achieved. While antiretroviral therapy (ART) has significantly improved disease outcomes for people with HIV-1 and reduced transmission, treatment is often accompanied by long-term side effects or stigma, or impeded by limited access to health care. Furthermore, because viral load quickly rebounds upon treatment interruption, ART is required to be a life-long medication. The major barrier to HIV cure is the persistence of long-lived latently infected CD4+ T cells. Collectively known as the latent reservoir, these cells carry integrated HIV-1 proviruses that are transcriptionally quiescent and are thus able to evade host immunity and virus-induced cell death. Cellular and molecular characterization of the latent reservoir is challenging because latently infected cells are exceedingly rare and express no known surface marker. Moreover, the infected cell pool is dominated by cells containing defective proviruses which cannot contribute to rebound viremia. The work herein elucidates the contribution of proviral integration site to HIV-1 latency and the maintenance of the replication-competent, or intact, reservoir. Using an innovative single-cell sequencing technique that provides paired proviral sequence and integration site information, as well as enables selective analysis of replication-competent HIV-1, I interrogate the integration landscape of HIV infected individuals whose reservoirs are dominated by a small number of large expanded clones. By performing viral outgrowth assays, I also show that replication-competent proviruses harbored in the expanded clones readily produce infectious virion upon stimulation. I report an integrative analysis of the clonal dynamics, inducibility, and genomic position of intact proviruses in ARTsuppressed individuals, and demonstrate that proviruses in expanded clones across patients are significant more likely to be mapped to Krüppel-associated box (KRAB) domain-containing zinc finger (ZNF) genes on chromosome 19. Transcriptional and epigenetic meta-analysis of primary CD4+ T cells reveal that these specific chromosomal locations harboring integrated provirus are associated with genes downregulated upon cellular activation. Taken together, the data indicate that selected sites in the genome, including ZNF genes, can be especially permissive for maintaining HIV-1 latency during memory CD4+ T cell clonal expansion. These findings demonstrate that gene activity at the integration site impacts the survival and persistence of intact, expanded HIV proviruses in infected cells, and provide evidence that the quality, not only the quantity, of the latent reservoir must a key consideration in HIV-1 cure strategies.

Comments

A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy

Recommended Citation

Huang, Amy S., "Integration Sites in the Persistence of Latent HIV-1" (2022). Student Theses and Dissertations. 667.
https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/667