Semester

Spring

Date of Graduation

2019

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Microbiology, Immunology, and Cell Biology

Committee Chair

Slawomir Lukomski

Committee Co-Chair

Christopher F. Cuff

Committee Member

Christopher F. Cuff

Committee Member

F. Heath Damron

Committee Member

Michael D. Schaller

Committee Member

Scott A. Weed

Abstract

Background: Group A Streptococcus (GAS) is responsible more than 700 million infections worldwide each year. Most of these infections start with initial colonization of the throat and skin, which is augmented by surface adhesins. The streptococcal collagen-like protein 1 (Scl1) is a major adhesin expressed by GAS that contains an N-terminal sequence-variable (V) domain, protruded away from the cell surface by the collagen domain. The Scl-V domain is comprised of three pairs of anti-parallel α-helices interconnected by surface-exposed loops. For attachment, GAS adhesins require a portal of entry, such as a wound or breach in the epithelium, to enter the body. Within the wound, host cells deposit a provisional extracellular matrix (ECM) rich in cellular fibronectin (cFn) isoforms that contain fibronectin type III (FnIII) repeats, including unique extra domains A (EDA) and B (EDB), that are absent in plasma fibronectin. Tenascin-C (TnC) is another ECM component, which is substantially deposited within the wound microenvironment and contains FnIII repeats. In addition to healing wounds, EDA/EDB-cFn and TnC are found in tumor microenvironments. Our early work showed that the Scl1-V domain binds to cFn via EDA. Our work here has focused on understanding this selective binding of Scl1 to wound-associated ECM and how this interaction contributes to GAS pathogenesis. Hypothesis: The Scl1-V domain has adapted the capability to bind to wound-associated FnIII repeats, EDA and EDB in cFn, and in TnC. Results: First, we discovered that surface-exposed loops of the Scl1-V domain, surrounding an acidic patch, participate in EDA binding. Both structural characteristics were conserved among phylogenetically distant Scl1 variants. We next discovered that Scl1 also binds to EDB in cFn, as well as to the FnIII repeats in TnC. Using defined recombinant proteins, we show that loop-region of the Scl1-V domain mediates Scl1 binding to EDB and to the TnC FnIII repeats. Moreover, Scl1-FnIII binding promotes GAS attachment and biofilm formation in vitro. We also developed a more complex heterogeneous in vitro matrix-system, deposited by cancer-associated fibroblasts, to demonstrate the selective binding of Scl1 to EDA/EDB-cFn isoforms and to TnC FnIII repeats within this matrix. Conclusions: Scl1 binds to wound-associated FnIII repeats in both cFn and TnC. This work has implications in GAS wound-colonization and infers Scl1-FnIII binding in modulating host wound-healing responses. This work lays a foundation for the development of strategies to target GAS during wound infections, as well as provides a rationale for the use of Scl1 in targeting cancerous tissues.

Embargo Reason

Publication Pending

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