Doctor of Philosophy, The Ohio State University, 2021, Microbiology

Viruses regulate host cell metabolism to promote infection. Fatty acid synthase (FASN) is a host enzyme that catalyzes palmitate synthesis by condensing acetyl and malonyl CoA. FASN is involved in the replication of the Human Immunodeficiency Virus-1 (HIV-1) and the Severe Acute Respiratory Syndrome Coronavirus type -2 (SARS-CoV- 2). FASN is found in the plasma of people living with HIV-1. The role of FASN in the extracellular milieu and its role in HIV-1 and coronavirus replication remain understudied. To study extracellular FASN during HIV-1 infection, we assayed FASN levels in longitudinal plasma samples from people with and without HIV-1. We found that people living with HIV-1 had eight- to ten-fold higher levels of plasma FASN relative to people without HIV-1. To understand FASN's role in the extracellular milieu, we exposed PBMCs to recombinant FASN and found increased secretion of TNF-α and IL-1β compared to exposure to the FASN vehicle control. If confirmed through follow-up studies, this would uncover a cytokine-like function for FASN. Fatty acids generated through FASN can be used for protein acylation, which is traditionally difficult to study. We developed a cell-permeable, click chemistry– compatible alkynyl acetate analog (Alk-4) that functions as a reporter of FASN dependent protein acylation. We showed that Alk-4 selectively labeled the cellular protein interferon-induced transmembrane protein 3 at its known palmitoylation sites and the HIV-1 matrix protein at its known myristoylation site in a FASN-dependent manner. Alk-4 also enabled biotin-based recovery of more than 200 FASN-dependent acylated cellular proteins. Alk-4 enabled the study of FASN's role in Gag myristoylation in HIV-1 infected cells, a process critical for membrane targeting of Gag. Using click chemistry and immunofluorescence techniques in Alk-4 labeled, HIV-1 infected cells as well as proximity ligation assays that produce a signal depending on the spat (open full item for complete abstract)

Committee: Jesse Kwiek (Advisor); Jacob Yount (Committee Member); Chad Rappleye (Committee Member); Karin Musier-Forsyth (Committee Member) Subjects: Microbiology; Virology