ABSTRACTEvident in its name, the gastric pathogenHelicobacter pylorihas a helical cell morphology which facilitates efficient colonization of the human stomach. An improved light-focusing strategy allowed us to robustly distinguish even subtle perturbations ofH. pyloricell morphology by deviations in light-scattering properties measured by flow cytometry. Profiling of an arrayed genome-wide deletion library identified 28 genes that influence different aspects of cell shape, including properties of the helix, cell length or width, cell filament formation, cell shape heterogeneity, and cell branching. Included in this mutant collection were two that failed to form any helical cells, a soluble lytic transglycosylase and a previously uncharacterized putative multipass inner membrane protein HPG27_0728, renamed Csd7. A combination of cell fractionation, mutational, and immunoprecipitation experiments show that Csd7 and Csd2 collaborate to stabilize the Csd1 peptidoglycan (PG) endopeptidase. Thus, bothcsd2andcsd7mutants show the same enhancement of PG tetra-pentapeptide cross-linking ascsd1mutants. Csd7 also links Csd1 with the bactofilin CcmA via protein-protein interactions. Although Csd1 is stable inccmAmutants, these mutants show altered PG tetra-pentapeptide cross-linking, suggesting that Csd7 may directly or indirectly activate as well as stabilize Csd1. These data begin to illuminate a highly orchestrated program to regulate PG modifications that promote helical shape, which includes nine nonessential nonredundant genes required for helical shape and 26 additional genes that further modifyH. pylori’s cell morphology.IMPORTANCEThe stomach ulcer and cancer-causing pathogenHelicobacter pylorihas a helical cell shape which facilitates stomach infection. Using light scattering to measure perturbations of cell morphology, we identified 28 genes that influence different aspects of cell shape. A mutant in a previously uncharacterized protein renamed Csd7 failed to form any helical cells. Biochemical analyses showed that Csd7 collaborates with other proteins to stabilize the cell wall-degrading enzyme Csd1. Csd7 also links Csd1 with a putative filament-forming protein via protein-protein interactions. These data suggest that helical cell shape arises from a highly orchestrated program to regulate cell wall modifications. Targeting of this helical cell shape-promoting program could offer new ways to block infectivity of this important human pathogen.
Newly Discovered Cell Shape Promoting Protein Complex Involved in the Maintenance of Distinct Helical Shape of Helicobacter Pylori
