Program Director/Principal Investigator (Last, First, Middle): Galn, Jorge PROJECT SUMMARY: Many symbiotic or pathogenic bacteria have evolved complex specialized machines known as type III secretion systems (T3SS), which have the capacity to transfer multiple bacterially-encoded proteins into host eukaryotic cells. These machines are of great interest because they are central to the pathogenic or symbiotic relationships of the bacteria that encode them. Proteins delivered by these machines, collectively referred to as ?effectors?, can modulate a great variety of eukaryotic cellular functions to shape the bacteria/host cell functional interphase. Through work supported by this Grant, we have been studying a T3SS from Salmonella enterica serovar Typhimurium (S. Typhimurium), encoded within its pathogenicity island 1 (SPI-1). This system mediates several phenotypes that are essential for virulence including bacterial entry into and survival within non-phagocytic cells, the induction of programmed cell death in macrophages, and the modulation of innate immune responses and inflammation in the intestinal tract. The central component of all T3SSs is the needle complex, a supramolecular structure which allows the translocation of proteins through the bacterial envelope during their journey to eukaryotic target cells. The needle complex works in association with other components such as the export apparatus, which mediates passage of the secreted proteins through the bacterial inner membrane, and the ?sorting platform?, a cytoplasmic structure that establishes an order in the secretion process. Additional components regulate the function of this machine to ensure that it is activated and deployed at the appropriate time (i. e. upon contact with target cells). Energy to drive protein translocation and the unfolding of the protein substrates is derived from an associated ATPase as well as a proton gradient established by poorly understood mechanisms. This research project intends to investigate less understood aspects of type III protein secretion including the mechanisms of sensing, signal transduction, and activation of the type III secretion machine, the deployment of the T3SS translocon into the target eukaryotic cell membrane, the structure and function of the T3SS export apparatus, and the mechanisms of by which the T3SS recognizes its substrates. Accomplishing these objectives will not only enhance our understanding of Salmonella spp. pathogenesis but also our understanding of T3SSs in general. Since this system is central to the pathogenesis of many important pathogenic bacteria, these studies may provide the bases for the development of broadly applicable anti-infective strategies.

Public Health Relevance

Many important bacterial pathogens such as Salmonella, Yersinia, Shigella, E. coli, Pseudomonas aeruginosa, Burkhodleria spp., Chlamydia spp., and Bordetella pertussis possess specialized nanomachine known as the type III protein secretion system, which is essential for their ability to cause disease. This machine ?injects? bacterial proteins into the cells of the host to alter cellular functions for the pathogen?s benefit. This project intends to study how this nanomachine works. The understanding of the mechanisms by which the type III secretion nanomachine works can lead to the development of broadly active anti infective strategies.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
4R37AI030492-32
Application #
9853860
Study Section
Special Emphasis Panel (NSS)
Program Officer
Alexander, William A
Project Start
1991-01-01
Project End
2025-06-30
Budget Start
2020-07-01
Budget End
2021-06-30
Support Year
32
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Yale University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520
Galán, Jorge E; Waksman, Gabriel (2018) Protein-Injection Machines in Bacteria. Cell 172:1306-1318
Park, Donghyun; Lara-Tejero, Maria; Waxham, M Neal et al. (2018) Visualization of the type III secretion mediated Salmonella-host cell interface using cryo-electron tomography. Elife 7:
Hu, Bo; Lara-Tejero, Maria; Kong, Qingke et al. (2017) In Situ Molecular Architecture of the Salmonella Type III Secretion Machine. Cell 168:1065-1074.e10
Zhang, Yongdeng; Lara-Tejero, María; Bewersdorf, Jörg et al. (2017) Visualization and characterization of individual type III protein secretion machines in live bacteria. Proc Natl Acad Sci U S A 114:6098-6103
Tsou, Lun K; Lara-Tejero, María; RoseFigura, Jordan et al. (2016) Antibacterial Flavonoids from Medicinal Plants Covalently Inactivate Type III Protein Secretion Substrates. J Am Chem Soc 138:2209-18
Zilkenat, Susann; Franz-Wachtel, Mirita; Stierhof, York-Dieter et al. (2016) Determination of the Stoichiometry of the Complete Bacterial Type III Secretion Needle Complex Using a Combined Quantitative Proteomic Approach. Mol Cell Proteomics 15:1598-609
Dietsche, Tobias; Tesfazgi Mebrhatu, Mehari; Brunner, Matthias J et al. (2016) Structural and Functional Characterization of the Bacterial Type III Secretion Export Apparatus. PLoS Pathog 12:e1006071
Monjarás Feria, Julia V; Lefebre, Matthew D; Stierhof, York-Dieter et al. (2015) Role of autocleavage in the function of a type III secretion specificity switch protein in Salmonella enterica serovar Typhimurium. MBio 6:e01459-15
Kato, Junya; Lefebre, Matthew; Galán, Jorge E (2015) Structural Features Reminiscent of ATP-Driven Protein Translocases Are Essential for the Function of a Type III Secretion-Associated ATPase. J Bacteriol 197:3007-14
Lefebre, Matthew D; Galán, Jorge E (2014) The inner rod protein controls substrate switching and needle length in a Salmonella type III secretion system. Proc Natl Acad Sci U S A 111:817-22

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