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Examination of the Role of HIV-1 Capsid Sequence on Virus Infectivity, Host Protein Interactions, and Capsid Uncoating

Fischer, Douglas Kite (2019) Examination of the Role of HIV-1 Capsid Sequence on Virus Infectivity, Host Protein Interactions, and Capsid Uncoating. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

Human immunodeficiency virus type 1 (HIV-1) infection persists for life, requiring the regular administration of antiretroviral therapy (ART) to suppress viral replication. Poorly tolerated side effects and the continued emergence of drug resistance mutations to current pharmaceuticals prompts the development of new therapies against novel targets. The HIV-1 capsid plays a critical role in every post-entry early virus life cycle step and its impairment is highly detrimental to successful infection. Combined with a high degree of mutational fragility and a lack of host cell analogs to either the monomeric capsid protein (CA) or the assembled capsid, these characteristics make capsid a viable target for therapeutic intervention. In this dissertation, we examined the influence of capsid from different virus strains on early virus life cycle steps in different cell types. We characterized differences in the infectivity defect of the cell cycle dependent CA mutation N57A when incorporated into the closely related lab-adapted virus strains HIV-1(NL4-3) and HIV-1(LAI). We extended these lab-adapted strain studies to include clinical isolates and demonstrated that a single CA amino acid polymorphism in the transmitted/founder strain HIV-1(CH040) enhances capsid stability compared with HIV-1(LAI). Examination of an HIV-1 sequence database revealed strong conservation of CA sequence and the potential for broader applicability of our findings in lab-adapted strains. Capsid interacts with numerous host proteins that facilitate virus infectivity. We examined the effects of inhibiting capsid interaction with three of these proteins, cleavage and polyadenylation specificity factor 6 (CPSF6), cyclophilin A (CypA), and nucleoporin 153 (Nup153), on virus infectivity. We also explored the manner in which a mislocalized, truncated form of CPSF6 (CPSF6-358) restricts virus infectivity. The capsid must become dissociated, or uncoat, within the host cell for early virus life cycle steps to proceed and lead to successful infection. We designed and performed initial testing of a new, live cell capable capsid permeabilization assay that will permit further insight into the initial steps of uncoating. Together, these studies serve to advance our understanding of how capsid functions to promote infection and how capsid mutations and differences in virus strains and cell types can alter virus infectivity.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Fischer, Douglas Kitedkf12@pitt.edudkf120000-0002-8267-6308
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorAmbrose, Zandreazaa4@pitt.edu
Committee MemberBomberger, Jenniferjbomb@pitt.edu
Committee MemberHoma, Fredflhoma@pitt.edu
Committee MemberSluis-Cremer, Nicolasnps2@pitt.edu
Committee MemberWatkins, Simonsimon.watkins@pitt.edu
Date: 25 May 2019
Date Type: Publication
Defense Date: 23 April 2019
Approval Date: 25 May 2019
Submission Date: 2 May 2019
Access Restriction: 3 year -- Restrict access to University of Pittsburgh for a period of 3 years.
Number of Pages: 317
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Microbiology and Immunology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: HIV-1 Capsid Uncoating Infectivity Mutation Sequence
Date Deposited: 25 May 2019 14:44
Last Modified: 25 May 2019 14:44
URI: http://d-scholarship.pitt.edu/id/eprint/36673

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