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- Title
Backbone Interactions Between Transcriptional Activator ExsA and Anti-Activator ExsD Facilitate Regulation of the Type III Secretion System in Pseudomonas aeruginosa.
- Authors
Shrestha, Manisha; Bernhards, Robert C.; Fu, Yichen; Ryan, Kylie; Schubot, Florian D.
- Abstract
The type III secretion system (T3SS) is a pivotal virulence mechanism of many Gram-negative bacteria. During infection, the syringe-like T3SS injects cytotoxic proteins directly into the eukaryotic host cell cytoplasm. In Pseudomonas aeruginosa, expression of the T3SS is regulated by a signaling cascade involving the proteins ExsA, ExsC, ExsD, and ExsE. The AraC-type transcription factor ExsA activates transcription of all T3SS-associated genes. Prior to host cell contact, ExsA is inhibited through direct binding of the anti-activator protein ExsD. Host cell contact triggers secretion of ExsE and sequestration of ExsD by ExsC to cause the release of ExsA. ExsA does not bind ExsD through the canonical ligand binding pocket of AraC-type proteins. Using site-directed mutagenesis and a specific in vitro transcription assay, we have now discovered that backbone interactions between the amino terminus of ExsD and the ExsA beta barrel constitute a pivotal part of the ExsD-ExsA interface. Follow-up bacterial two-hybrid experiments suggest additional contacts create an even larger protein–protein interface. The discovered role of the amino terminus of ExsD in ExsA binding explains how ExsC might relieve the ExsD-mediated inhibition of T3SS gene expression, because the same region of ExsD interacts with ExsC following host cell contact.
- Subjects
PSEUDOMONAS aeruginosa; GRAM-negative bacteria; GENE expression; TRANSCRIPTION factors; CELLULAR signal transduction
- Publication
Scientific Reports, 2020, Vol 10, Issue 1, p1
- ISSN
2045-2322
- Publication type
Article
- DOI
10.1038/s41598-020-66555-z