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The role of fatty acid synthase in viral replication

Karthigeyan, Krithika Priyadarshini
http://orcid.org/0000-0002-1808-6688
http://rave.ohiolink.edu/etdc/view?acc_num=osu1639680004140588

Abstract Details

2021, Doctor of Philosophy, Ohio State University, 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 spatial distance between two interacting entities in a cell, we showed co-localization of Alk-4 adducts with HIV-1 Gag. Treatment with Fasnall, a selective and potent FASN inhibitor, reduced the colocalization of Alk-4 adducts with Gag. To identify if exogenous or de novo sources of myristate are preferentially used for Gag myristoylation, we developed a dual-click chemistry approach using analogs of exogenous and de novo myristate. While the exogenous myristate source labeling of HIV-1 MA was stronger, this might be a result of excess concentrations of exogenous myristate in our assay compared to physiological levels. As FASN is involved in coronavirus replication, we tested Fasnall against bovine coronavirus and SARS-CoV-2. Fasnall inhibited the replication of both viruses. Fasnall also reduced SARS-CoV-2 Spike expression in transfected cells. As the SARS-CoV-2 Spike protein is palmitoylated, labeling of multiple Spike constructs with Alk-4 revealed site-specific labeling of the Spike protein. In conclusion, FASN plays a role in the replication of HIV-1 and SARS-CoV-2. There is a need for broad-spectrum antivirals that safely and potently block replication of several viruses. FASN can be an ideal pan antiviral target due to its limited expression in healthy adult cells, viability of tissue specific FASN knockouts and its requirement for the replication of multiple viruses.
Jesse Kwiek (Advisor)
Jacob Yount (Committee Member)
Chad Rappleye (Committee Member)
Karin Musier-Forsyth (Committee Member)
218 p.
Microbiology; Virology
Fatty acid synthase; HIV-1; SARS-CoV-2; Influenza; Bovine coronavirus, click chemistry, S-palmitoylation; N-myristoylation; fatty acylation; IFITM3; HIV-1 Gag; Fasnall

Recommended Citations

Citations

  • Karthigeyan, K. P. (2021). The role of fatty acid synthase in viral replication [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1639680004140588

    APA Style (7th edition)

  • Karthigeyan, Krithika. The role of fatty acid synthase in viral replication. 2021. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1639680004140588.

    MLA Style (8th edition)

  • Karthigeyan, Krithika. "The role of fatty acid synthase in viral replication." Doctoral dissertation, Ohio State University, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=osu1639680004140588

    Chicago Manual of Style (17th edition)

osu1639680004140588
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This open access ETD is published by The Ohio State University and OhioLINK.