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Glycosaminoglycan binding of strain-specific polymorphisms in the chikungunya virus E2 glycoprotein

Graudin, Grace Winesett (2024) Glycosaminoglycan binding of strain-specific polymorphisms in the chikungunya virus E2 glycoprotein. Master's Thesis, University of Pittsburgh. (Unpublished)

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Abstract

Chikungunya virus (CHIKV) is an arthritogenic alphavirus that displays broad cell tropism, interacting with a variety of cell-surface molecules to enter cells and mediate infection. Like many pathogenic viruses, CHIKV uses negatively-charged glycosaminoglycans (GAGs) as attachment factors to enhance binding to cells. Multiple lines of evidence suggest that CHIKV depends on GAGs for efficient infection of mammalian cells and displays strain-specific differences in GAG binding. However, the specific polymorphisms that dictate strain-specific differences in GAG binding have not been identified. We engineered SINV-CHIKV chimeric viruses to contain five basic amino acid polymorphisms at residues in the E2 viral attachment protein that either have putative roles in GAG binding or are present in strains displaying phenotypic differences (K140R, K149R, K221R, K234N, and K252Q). ELISA assays were conducted to determine whether the E2 polymorphic mutants were altered in direct binding to GAGs while cell-binding assays were conducted using human muscle cells (RH30) to determine whether these mutants depend on GAGs for efficient cell attachment. In addition, mutant viruses were also tested in infectivity assays to investigate their dependence on GAGs for infection of biologically-relevant cell lines (RH30, pHDF, tel-HFF) and dependence on GAGs for efficient infection in the presence and absence of a CHIKV entry receptor (Mxra8). Collectively, these studies provide a foundational understanding of the functional consequences of CHIKV E2 basic amino acid polymorphisms in binding to GAGs and cell attachment.

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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Graudin, Grace Winesettgrw59@pitt.edugrw59
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorSilva, Laurielaurie.silva@pitt.edulaurie.silva
Committee MemberMattila, Joshuajmattila@pitt.edujmattila
Committee MemberMcMillen, Cynthiacmm287@pitt.educmm287
Committee MemberMcElroy, Anitamcelroya@pitt.edumcelroya
Date: 14 May 2024
Date Type: Publication
Defense Date: 16 April 2024
Approval Date: 14 May 2024
Submission Date: 22 April 2024
Access Restriction: 2 year -- Restrict access to University of Pittsburgh for a period of 2 years.
Number of Pages: 87
Institution: University of Pittsburgh
Schools and Programs: School of Public Health > Infectious Diseases and Microbiology
Degree: MS - Master of Science
Thesis Type: Master's Thesis
Refereed: Yes
Uncontrolled Keywords: Chikungunya virus, arbovirus, glycosaminoglycans, viral attachment, viral entry, virus-host interactions
Date Deposited: 14 May 2024 19:22
Last Modified: 14 May 2024 19:22
URI: http://d-scholarship.pitt.edu/id/eprint/46232

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