Group A Streptococcus (GAS), despite being among the best-studied human pathogens, continues to cause significant morbidity and mortality. The pathogen can result in invasive infections, such as necrotizing fasciitis or streptococcal toxic shock, in addition to the more common oropharyngeal or skin infections. The CsrRS two-component signaling system of GAS is thought to play a major role in virulence and regulates 10-15% of the pathogen's genome. Recently, it was shown that LL-37, a human antimicrobial peptide, can paradoxically lead to increased expression of virulence factors through this two-component system. We have evidence for a direct binding interaction between LL-37 and the extracellular domain of the CsrS receptor. We have also demonstrated that the receptor is able to dimerize independently of the intracellular domain. The goals of this study are to extend these observations and to determine the binding site for LL-37 on CsrS. We will also determine possible ligand-induced conformational changes and the role of the dimer conformation on CsrS signaling. We will determine the structure of the CsrS extracellular domain with its transmembrane domains embedded in a membrane nanodisc and expand these techniques to an additional two-component receptor. These data will lead to a better understanding of the structure and function of CsrS signaling by LL-37 and will establish the foundation for continued studies of GAS signaling mechanisms and potential therapeutic inhibition, and signaling by two-component receptors in general. As part of the proposed training program, one of the short-term goals is to acquire skills in the study of bacterial signaling systems and the biochemistry and structural biology of membrane proteins. The project will be developed under the mentorship of Dr. Michael Wessels, a leader and expert in the field of GAS pathogenesis, and Dr. James Chou, an expert in the structural biology of membrane proteins; a scientific advisory committee composed of experts in bacterial pathogenesis and structural biology will provide advice on overall direction of the project and career guidance. The training program will include coursework in structural biology and crystallography, membrane biochemistry, and computational biology; and didactic learning from local seminars as well as national and international conferences. The research will take place in the laboratories of Dr. Wessels at Boston Children's Hospital and Dr. Chou at Harvard Medical School. The Harvard medical campus is a dynamic research environment with multiple opportunities for professional development and expertise in all aspects of the proposed research. Boston Children's Hospital is also an extremely supportive environment and is committed to providing 85% protected time for this research. The project will build on the candidate's past experiences in microbial pathogenesis and NMR spectroscopy and allow him to pursue a career as a physician-scientist with the goal of an academic career in a pediatric division of infectious diseases.
Invasive group A streptococcal infections such as necrotizing fasciitis and streptococcal toxic shock cause significant morbidity and mortality. The pathogen uses a signaling system (the CsrRS two-component system) to regulate a number of virulence factors. This study seeks to understand the mechanism by which the receptor in this system responds to an environmental signal and leads to upregulation of virulence factors, and will eventually contribute to the development of possible therapeutics aimed at inhibiting bacterial signaling mechanisms.
|Velarde, Jorge J; O'Seaghdha, Maghnus; Baddal, Buket et al. (2017) Binding of NAD+-Glycohydrolase to Streptolysin O Stabilizes Both Toxins and Promotes Virulence of Group A Streptococcus. MBio 8:|