Molecular analysis of Staphylococcus aureus toxin interaction with the host /

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Bibliographic Details
Author / Creator:Berube, Bryan Joshua, author.
Ann Arbor : ProQuest Dissertations & Theses, 2015
Description:1 electronic resource (155 pages)
Format: E-Resource Dissertations
Local Note:School code: 0330
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Other authors / contributors:University of Chicago. degree granting institution.
Notes:Advisors: Juliane Bubeck Wardenburg Committee members: Tatyana Golovkina; Olaf Schneewind; Howard Shuman; Jerrold Turner.
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Dissertation Abstracts International, Volume: 76-11(E), Section: B.
Summary:Staphylococcus aureus is a major human pathogen that causes a wide spectrum of disease manifestations ranging from relatively mild skin and soft tissue infections to life-threatening diseases, such as pneumonia and sepsis. Traditionally considered an opportunistic healthcare-associated pathogen, the last few decades have seen an increase in highly virulent antibiotic-resistant strains capable of causing disease in otherwise healthy individuals. These attributes have led to increased morbidity and mortality, as well as a large economic burden on the healthcare system.
The success of S. aureus stems from its ability to elaborate a broad range of virulence factors, including numerous secreted toxins. Studies of these virulence factors are critical to understanding S. aureus disease pathogenesis and for the development of novel preventatives and therapeutics. Two secreted toxins that have been extensively studied for their contribution to S. aureus disease are the pore-forming cytolysin, alpha-toxin (Hla) and the amphipathic ?-helical phenol-soluble modulin (PSM) peptides. Our work elucidates novel molecular mechanisms and cell biological outcomes of Hla intoxication, while also describing the integration of Hla with PSMs during disease pathogenesis.
Utilizing a eukaryotic expression system, we show the first gain-of-function study to confirm A Disintegrin And Metalloprotease (ADAM10) as the cellular receptor for Hla. We use this system to highlight the metalloprotease and cysteine-rich domains of ADAM10 as the binding sites for Hla and provide the first demonstration of dissociation between toxin pore formation and upregulation of ADAM10 enzymatic activity. Next, we highlight the host cell response following Hla binding to show that toxin pore formation induces the endocytosis and aggregation of the toxin-receptor complex into early endosomes. We show that endocytosis of Hla is host-cell protective in that it diminishes both toxin-mediated cytotoxicity and upregulation of ADAM10 activity. Following internalization, Hla alters the normal trafficking pathway, blocking endosomal acidification and causing the formation of enlarged early endosomes, the consequence of which remains unclear. Finally, we utilize genetic mutants in the hla and psm loci to demonstrate the integrated activity of the two toxins. We show that the psmalpha locus regulates production of Hla and causes a defect in Hla secretion, which is responsible for the virulence defect in pneumonia and skin and soft tissue models of infection. The Hla defect can be overcome by plasmid-driven overexpression of Hla, which restores virulence to the S. aureus psm mutant. Regulation of Hla production by PSMs ensures coordinated secretion of the two toxins. We show that PSMs cause synergistic lysis of host cells in the presence of Hla, likely through signaling through its receptor, formyl peptide receptor 2. Together, our results highlight novel aspects of Hla intoxication and begin to study the integration of multiple virulence factors during disease, which will be critical when designing vaccines to prevent staphylococcal disease.