The type III secretion determinants of the flagellar anti‐transcription factor, FlgM, extend from the amino‐terminus into the anti‐σ28 domain

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Abstract
The flagellar‐specific anti‐sigma factor, FlgM, inhibits the expression of late flagellar genes until the hook–basal body structure is assembled and competent for export of the flagellins and hook‐associated proteins (flagellar late proteins). FlgM monitors this assembly checkpoint by being a substrate for export via the hook–basal body structure, which includes a type III protein secretion complex. Amino acid sequence alignment of late‐secreted flagellar proteins identified a region of homology present in the amino‐terminus of FlgM and the other late flagellar proteins, but not in flagellar proteins secreted earlier during flagellar biosynthesis. Single amino acid substitutions at specific positions within this motif decreased the export of FlgM. Deletion of this region (S3‐P11) resulted in lower intracellular FlgM levels, but did not prevent recognition and export by the flagellar‐specific secretion system. Mutations were isolated in a second region of FlgM spanning residues K27 to A65 that exhibited increased anti‐σ28 activity. These FlgM ‘hyperinhibitor’ mutants were secreted less than wild‐type FlgM. Mutations that interfere with the secretion of FlgM without abolishing anti‐σ28 activity have a negative effect upon the secretion of a His‐tagged FlgM mutant that lacks anti‐σ28 activity. Models are proposed to explain the dominant negative phenotype of the FlgM secretion mutants reported in this study.