Dissecting bacterial secretion systems in situ with electron cryo-tomography
EMSL Project ID
50989
Abstract
Pathogenic bacteria use specialized “nanomachines” to identify and interact with host cells. These machines are attractive drug targets because they are surface-exposed, widespread, and vital for pathogenicity. While one of these machines, the Type III Secretion System, has been well studied, other systems remain relatively poorly understood. Here, we propose to use electron cryotomography (cryo-ET) to dissect the structures and functions of multiple pathogenic nanomachines. Previously, we applied cryo-ET to the Type VI Secretion System, where our images immediately revealed its "spring-loaded" contractile mechanism. Going a step further, we realized that we could combine cryo-ET and high-resolution subtomogram averaging with available knowledge from other techniques to produce a complete architectural model of the Myxococcus xanthus Type IVa Pilus (a Type II Secretion System). This produced a flood of new mechanistic insights and inspired us to apply the same approach to pathogenic secretion systems. In this project, we propose to use cryo-ET to reveal the structure and function of Type II, Type III, Type IV, Type V, Type VI, Type VII and Type IX secretion systems, combining subtomogram averaging with difference analysis of mutants in which individual components are knocked out or tagged in order to produce architectural models of the structures. By comparing the structures of pathogenic and non-pathogenic relatives, we aim to identify adaptations underlying virulence functions. Together, we expect this project to produce a detailed mechanistic picture of the nanomachines that mediate pathogenesis, an important first step in identifying therapeutic targets in the future. To accomplish these goals, we request programmatic access at PNCC (totaling 20 days per year).
Project Details
Start Date
2019-09-01
End Date
2021-03-17
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members