Chlamydia Pneumoniae CdsL Regulates CdsN ATPase Activity, and Disruption with a Peptide Mimetic Prevents Bacterial Invasion
Authors: Elisa A Porfilio
Jerry W Slootstra
James B Mahony
David C Bulir
Connor A Emdin
Ryan M Pirie
Chris B Stone
Journal: Frontiers in Microbiology (2011)
Discipline: MICROBIOLOGY
Keywords: ATPase; Chlamydia; Pepscan; peptide mimetic; type III secretion.
Affiliations: Michael G. DeGroote Institute for Infectious Disease Research, Faculty of Health Sciences, Department of Pathology and Molecular Medicine, McMaster University, Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare Hamilton, ON, Canada.

Abstract:  Chlamydiae are obligate intracellular pathogens that likely require type III secretion (T3S) to invade cells and replicate intracellularly within a cytoplasmic vacuole called an inclusion body. Chlamydia pneumoniae possess a YscL ortholog, CdsL, that has been shown to interact with the T3S ATPase (CdsN). In this report we demonstrate that CdsL down-regulates CdsN enzymatic activity in a dose-dependent manner. Using Pepscan epitope mapping we identified two separate binding domains to which CdsL binds viz. CdsN(221-229) and CdsN(265-270). We confirmed the binding domains using a pull-down assay and showed that GST-CdsN(221-270), which encompasses these peptides, co-purified with His-CdsL. Next, we used orthology modeling based on the crystal structure of a T3S ATPase ortholog from Escherichia coli, EscN, to map the binding domains on the predicted 3D structure of CdsN. The CdsL binding domains mapped to the catalytic domain of the ATPase, one in the central channel of the ATPase hexamer and one on the outer face. Since peptide mimetics have been used to disrupt essential protein interactions of the chlamydial T3S system and inhibit T3S-mediated invasion of HeLa cells, we hypothesized that if CdsL-CdsN binding is essential for regulating T3S then a CdsN peptide mimetic could be used to potentially block T3S and chlamydial invasion. Treatment of elementary body with a CdsN peptide mimetic inhibited C. pneumoniae invasion into HeLa cells in a dose-dependent fashion. This report represents the first use of Pepscan technology to identify binding domains for specific T3S proteins viz. CdsL on the ATPase, CdsN, and demonstrates that peptide mimetics can be used as anti-virulence factors to block bacterial invasion. 

Retracted: No

PubPeer comments:
 Figure 5. * The figure legend descriptions for A (dose-response) and B (time series) appear to have been switched. Could the authors please check? * Red boxes: The a-GST and Loading Control blots in panel B appear to look very similar. The use of the same LC in panel A is appropriate and not of concern. * Green arrows: Some of the a-GST bands appear to be surrounded by sharp background transitions, as if these samples were not all run next to each other. The corresponding loading control does not show these transitions. Could the authors please provide the original blots? ![file] , 
 Correction, 12 October 2021 found here: https: www.frontiersin.org articles 10.3389 fmicb.2021.726518 full The correction reads:  In the original article, there was a mistake in Figure 5 as published. Two lanes of a gel were fused together to decrease the size of the figure and improve clarity, when in fact they were not run in side-by-side lanes on the gel. The corrected Figure 5 appears below. [...] The authors apologize for this error. Given this correction to Figure 5, internalization of the peptide is now unclear and we do not have strong evidence to suggest that the peptide is affecting an intracellular protein interaction. Further studies are required to support this hypothesis. The original article has been updated. Could the authors please clarify why they never replied here, and how this updated finding changes the conclusions of the paper? ,