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VacA generates a protective intracellular reservoir for Helicobacter pylori that is eliminated by activation of the lysosomal calcium channel TRPML1

Abstract

Helicobacter pylori infection is a proven carcinogen for gastric cancer. Its virulence factor vacuolating cytotoxin A (VacA) promotes more severe disease and gastric colonization. VacA, by an unknown mechanism, usurps lysosomal and autophagy pathways to generate a protected reservoir for H. pylori that confers bacterial survival in vitro. Here, we show the existence of a VacA-generated intracellular niche in vivo that protects the bacteria from antibiotic treatment and leads to infection recrudescence after therapy. Furthermore, we report that VacA targets the lysosomal calcium channel TRPML1 to disrupt endolysosomal trafficking and mediate these effects. Remarkably, H. pylori that lack toxigenic VacA colonize enlarged dysfunctional lysosomes in the gastric epithelium of trpml1-null mice, where they are protected from eradication therapy. Furthermore, a small molecule agonist directed against TRPML1 reversed the toxic effects of VacA on endolysosomal trafficking, culminating in the clearance of intracellular bacteria. These results suggest that TRPML1 may represent a therapeutic target for chronic H. pylori infection.

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Fig. 1: VacA generates an intracellular reservoir in vivo leading to bacterial persistence after eradication therapy.
Fig. 2: VacA impairs TRPML1 activity.
Fig. 3: TRPML1 deficiency generates a VacA-like intracellular niche for H. pylori in vivo.
Fig. 4: TRPML1 activation restores VacA-disrupted endolysosomal and autophagy pathways.
Fig. 5: Validation of the VacA–TRPML1 axis in human gastric organoids.
Fig. 6: TRPML1 activation eliminates both the intracellular niche and survival advantage of VacA+ H. pylori.

Data availability

Material used in this study is readily available from the authors or from the commercial source. AGS cells and H. pylori 60190 strain are available from the corresponding author.

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Acknowledgements

We thank S. Slaugenhaupt for the Trpml1-deficient mice, S. Blanke for the purified VacA toxin and anti-VacA polyclonal antibody and J. Atherton for the H. pylori SS1 strains. This work was supported by the Canadian Institutes of Health Research (CIHR), the Canadian Association of Gastroenterology (CAG) and the North American Society for Paediatric Gastroenterology, Hepatology and Nutrition Foundation. L.K.G. was supported by a CIHR/CAG/Canadian Crohn’s and Colitis and CIHR/CAG/AbbVie Pharmaceuticals Canada Fellowship.

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Authors

Contributions

N.L.J. conceived and supervised the study, analysed the data and wrote the manuscript. M.I.C., L.K.G. and A.P. designed, carried out and analysed most of the experiments and wrote the manuscript. S.X., M.A., N.B.-S., J.C., Y.Z., C.O’B. and M.H. assisted with the gastric organoid studies and provided helpful comments for the manuscript. X.Z.Z. and X.D. performed the lysosomal calcium release experiments, analysed the data and provided helpful comments for the manuscript. R.P. provided the human biopsies and I.S. analysed the H. pylori staining and both provided helpful comments for the manuscript. H.W., L.R. and D.J.P. contributed to experimental design and provided helpful comments for the manuscript.

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Correspondence to Nicola L. Jones.

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Supplementary information

Supplementary Information

Supplementary Table 1, Supplementary Figures 1–19, uncropped blots and Supplementary Video legends.

Reporting Summary

Supplementary Video 1

SS1 H. pylori is restricted to the glandular lumen in wild-type mice. H. pylori staining of gastric tissue obtained from wild-type infected mice. The video was created by a 3D reconstruction of confocal z-sections acquired each 0.20 µm using a 40× water objective and deconvolved utilizing Volocity software. Similar staining was obtained for all of the wild-type mice (n = 12).

Supplementary Video 2

SS1 H. pylori colonize vacuolar compartments of parietal cells in trpml1−/− mice. H. pylori staining of gastric tissue obtained from Trpml1-deficient infected mice. The video was created by a 3D reconstruction of confocal z-sections acquired each 0.20 µm using a 40× water objective and deconvolved utilizing Volocity software. H. pylori within vacuolar compartments in parietal cells were identified in all of the Trpml1-deficient mice analysed (n = 8).

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Capurro, M.I., Greenfield, L.K., Prashar, A. et al. VacA generates a protective intracellular reservoir for Helicobacter pylori that is eliminated by activation of the lysosomal calcium channel TRPML1. Nat Microbiol 4, 1411–1423 (2019). https://doi.org/10.1038/s41564-019-0441-6

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