ORCID

http://orcid.org/0000-0002-3398-9316

Date of Award

Winter 1-15-2021

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Molecular Microbiology & Microbial Pathogenesis)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

Human immunodeficiency virus type 1 (HIV-1) relies on a handful of essential enzymes for replication. Among these, the viral integrase enzyme (IN) plays a pivotal role in the viral life cycle by catalyzing the integration of the reverse-transcribed viral DNA into the host chromosome. While integration is the canonical role of IN, new research has uncovered an additional vital role for IN during virion morphogenesis. This dissertation elucidates how IN contributes to proper packaging of the viral RNA genome (vRNA) within the viral capsid and examines the fate of improperly formed viral particles in target cells.IN is proposed to mediate proper placement of the vRNA within the capsid in mature virions by binding to vRNA at a defined binding site in its C-terminal domain. Mutations at the CTD RNA-binding site lead to the generation of morphologically aberrant virions with vRNA mislocalized outside of the empty capsid lattice, suggesting that IN-RNA binding is necessary for proper virion maturation. However, multiple IN mutations outside of the RNA-binding site cause the same morphological defects in virions, suggesting that another property of IN, such as its multimerization, may be responsible for its role in virion morphogenesis. In Chapter 2 we dissect the contribution of IN-RNA binding and IN multimerization to virion morphogenesis and demonstrate that IN-RNA binding accounts for the role of IN in virion maturation, although IN tetramerization is likely a prerequisite for RNA binding to occur. We further identify three separate mechanisms by which IN-RNA interaction can be inhibited, all of which prevent proper virion maturation. In Chapter 3, we provide evidence that the morphological defects caused by IN mutations lead to the premature loss of the exposed vRNA and IN itself in infected target cells, preventing further viral replication. As a whole, this dissertation provides mechanistic insight into how IN contributes to virion morphogenesis by establishing IN-RNA binding as the determining factor by which IN ensures proper placement of the vRNA in viral particles, while highlighting the importance of proper IN multimerization for RNA-binding. This work also provides an explanation for the common block in viral replication observed in many IN mutant viruses.

Language

English (en)

Chair and Committee

Sebla B. Kutluay

Committee Members

Deborah J. Lenschow, Michael S. Diamond, Jacco Boon, Gwendalyn J. Randolph,

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Virology Commons

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