Date of Award

Summer 8-23-2025

Document Type

Dissertation

Degree Name

Ph.D. in Biological Sciences

Organizational Unit

College of Natural Science and Mathematics, Biological Sciences

First Advisor

Schuyler B. van Engelenburg

Second Advisor

Todd Blankenship

Third Advisor

Dinah Loerke

Fourth Advisor

Kingshuk Ghosh

Copyright Statement / License for Reuse

All Rights Reserved
All Rights Reserved.

Keywords

AIDS, Biochemistry, Cellular biology, HIV-1, Life sciences, Molecular biology

Abstract

Human Immunodeficiency Virus type-1 (HIV-1) infects CD4+ T cells and remains a global pandemic due to our incomplete understanding of the molecular mechanisms underlying its highly coordinated and regulated assembly process and the strategies it uses for immune evasion. For HIV-1 to produce infectious particles, the viral envelope glycoprotein (Env) must be selectively trapped and incorporated into assembly sites formed by the Gag polyprotein multimerizing at the plasma membrane. This process is regulated by the long cytoplasmic tail of Env (EnvCT) in concert with the immature Gag lattice. However, the mechanisms of this regulatory engagement and the broader mechanisms by which HIV-1 controls assembly remain incompletely understood.

This work addresses three major aspects of Env incorporation: the trafficking fate of unincorporated Env, the quaternary state of the EnvCT, and how Gag lattice organization influences incorporation.

Using a biochemical pulse-chase assay, I found that internalized Env traffics to Rab14+ compartments with features of late endosomes and lysosomes. These findings clarify the post-endocytic fate of surface Env and provide insight into how surface levels are regulated in infected T cells.

To determine whether the quaternary structure of the EnvCT trimer is required for incorporation, I assessed monomeric EnvCT chimeras using a domain-specific bivalent crosslinking reagent. These constructs remained monomeric in both cellular and virion contexts, yet were still incorporated into virus-like particles. This demonstrates that a single EnvCT is sufficient for lattice trapping and incorporation.

Structural defects in the Gag lattice may serve as sites of Env incorporation, but this possibility had not been previously tested. I introduced mutations within Gag that increase lattice defects and observed altered Env incorporation across multiple T cell lines, suggesting that lattice accessibility and spatial arrangement can influence the number or availability of incorporation sites.

Altogether, this work advances our understanding of how HIV-1 selectively incorporates Env through coordinated regulation of trafficking, cytoplasmic tail structure, and Gag lattice architecture.

Copyright Date

8-2025

Publication Statement

Copyright is held by the author. User is responsible for all copyright compliance.

Rights Holder

Austin Robert Clark

Provenance

Received from ProQuest

File Format

application/pdf

Language

English (eng)

Extent

131 pgs

File Size

13 MB

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Available for download on Saturday, September 18, 2027



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