Multiple Peptidoglycan Modification Networks Modulate Helicobacter pylori's Cell Shape, Motility, and Colonization Potential

Abstract

Helical cell shape of the gastric pathogen Helicobacter pylori has been suggested to promote virulence through viscosity-dependent enhancement of swimming velocity. However, H. pylori csd1 mutants, which are curved but lack helical twist, show normal velocity in viscous polymer solutions and the reason for their deficiency in stomach colonization has remained unclear. Characterization of new rod shaped mutants identified Csd4, a DL-carboxypeptidase of peptidoglycan (PG) tripeptide monomers and Csd5, a putative scaffolding protein. Morphological and biochemical studies indicated Csd4 tripeptide cleavage and Csd1 crosslinking relaxation modify the PG sacculus through independent networks that coordinately generate helical shape. csd4 mutants show attenuation of stomach colonization, but no change in proinflammatory cytokine induction, despite four-fold higher levels of Nod1-agonist tripeptides in the PG sacculus. Motility analysis of similarly shaped mutants bearing distinct alterations in PG modifications revealed deficits associated with shape, but only in gel-like media and not viscous solutions. As gastric mucus displays viscoelastic gel-like properties, our results suggest enhanced penetration of the mucus barrier underlies the fitness advantage conferred by H. pylori's characteristic shape.

Author Summary

The only habitat of Helicobacter pylori is the human stomach, where it can promote stomach ulcers and cancer. Cells lining the stomach are protected from luminal acid by a thick layer of gastric mucus composed of polymerized gastric mucins. Gastric mucin undergoes a physical transition between a viscoelastic solution at neutral pH to a viscoelastic gel-like state at low pH. Helical rod shape in bacteria has been suggested to enhance swimming velocity in viscous solutions by a cork-screw mechanism, but H. pylori mutants lacking helical twist show normal swimming velocity in viscous polymer solutions used in prior studies comparing motility across bacterial species. These same mutants, however, show diminished colonization suggesting helical shape promotes stomach infection by another mechanism. Here we identified Csd4, a protease of cell wall tripeptides, which induces curvature in the cell body independently from the changes in cell wall crosslinking previously shown to promote helical twist. Cells lacking Csd4 form straight rods that also show colonization defects but normal velocity in several viscous polymer solutions. Upon examination of motility in gel-like media, however, we discovered that elimination or exaggeration of cell curvature perturbs motility. Thus H. pylori's helical shape may aid penetration of gel-like stomach mucus.