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Non-canonical roles of Oligoadenylate Synthetase 1 during viral and bacterial infections

Perez, Joseph (2020) Non-canonical roles of Oligoadenylate Synthetase 1 during viral and bacterial infections. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

Interferons inhibit growth of several intracellular pathogens, including virus and bacteria, through the expression of interferon stimulated genes (ISGs). We have found that a specific isoform of one such ISG, Oligoadenylate Synthetase 1 (OAS1) enhances translation of a select set of mRNAs, thereby increases the steady state and induced levels of specific proteins with antiviral and antibacterial properties. This OAS1 isoform (OAS1 P46) in humans is generated due to an alternative splice acceptor site at the C-terminus of OAS1 gene. The SNP rs10774671 at this site has been associated with disease severity to West Nile Virus (WNV) and Mycobacterium tuberculosis. We show that human OAS1-KO cells have lower basal levels of cGAS protein and can be rescued by OAS1 P46 expression, independent of its enzyme activity. Inducible expression of OAS1 P46 in cGAS-KO cells does not suppress WNV replication, suggesting that the antiviral activity of OAS1 is mediated through cGAS. We also have established functional equivalence between OAS1 P46 and a mouse ortholog, Oas1b (no enzyme activity), which similarly affects WNV susceptibility. Through RNA-protein crosslinking experiments we have identified target mRNAs that bind to OAS1 and Oas1b. We have demonstrated increased sensitivity of WNV in OAS1/Oas1b RNA binding mutants. To define the antibacterial activity of OAS1, we screened OAS1-deficient THP1 cells for bacterial growth using the intracellular bacteria Listeria monocytogenes and Francisella novicida as model bacterial pathogens. Inducible expression of OAS1 P46 WT and enzymatically inactive mutant rescued this antibacterial activity of OAS1 in OAS1-KO cells. Further investigation of OAS1-KO cells indicated defective type II IFN signaling due to the reduced expression of IRF1. Like cGAS, OAS1 bound to IRF1 mRNA and enhanced protein synthesis of IRF1. Inducible expression of OAS1 P46 in IRF1-KO cells did not inhibit bacterial growth suggesting that antibacterial activity of OAS1 is mediated through IRF1. Lastly, we found that Oas1b knock-in (Oas1b-KI) mice showed improved survival with L. monocytogenes or F. novicida as compared to WT mice. These findings suggest a new mechanism of OAS1 in which it binds to target mRNAs, enhances the translation of these RNAs and limits virus and bacterial infections.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Perez, Josephjop84@pitt.edujop84
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorSarkar, Saumendra / Nsaumen@pitt.edu
Committee ChairDeLuca, Neal / Andeluca@pitt.edu
Committee MemberBrodsky, Jeffreyjbrodsky@pitt.edu
Committee MemberDelgoffe, Greg / Mdelgoffeg@upmc.edu
Committee MemberWilliams, John / Vjvw@pitt.edu
Date: 24 August 2020
Date Type: Publication
Defense Date: 3 August 2020
Approval Date: 24 August 2020
Submission Date: 7 August 2020
Access Restriction: 2 year -- Restrict access to University of Pittsburgh for a period of 2 years.
Number of Pages: 213
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: Virology, Bacteriology, Innate Immunity, OAS
Date Deposited: 24 Aug 2020 04:00
Last Modified: 15 Sep 2020 19:59
URI: http://d-scholarship.pitt.edu/id/eprint/39577

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