The Serine Protease Autotransporter Pic Modulates Citrobacter rodentium Pathogenesis and Its Innate Recognition by the Host

Bacterial pathogens produce a number of autotransporters that possess diverse functions. These include the family of serine protease autotransporters of Enterobacteriaceae (SPATEs) produced by enteric pathogens such as Shigella flexneri and enteroaggregative Escherichia coli. Of these SPATEs, one termed "protein involved in colonization," or Pic, has been shown to possess mucinase activity in vitro, but to date, its role in in vivo enteric pathogenesis is unknown. Testing a pic null (Delta picC) mutant in Citrobacter rodentium, a natural mouse pathogen, found that the C. rodentium Delta picC strain was impaired in its ability to degrade mucin in vitro compared to the wild type. Upon infection of mice, the Delta picC mutant exhibited a hypervirulent phenotype with dramatically heavier pathogen burdens found in intestinal crypts. Delta picC mutant-infected mice suffered greater barrier disruption and more severe colitis and weight loss, necessitating their euthanization between 10 and 14 days postinfection. Notably, the virulence of the Delta picC mutant was normalized when the picC gene was restored; however, a PicC point mutant causing loss of mucinase activity did not replicate the Delta picC phenotype. Exploring other aspects of PicC function, the Delta picC mutant was found to aggregate to higher levels in vivo than wild-type C. rodentium. Moreover, unlike the wild type, the C. rodentium Delta picC mutant had a red, dry, and rough (RDAR) morphology in vitro and showed increased activation of the innate receptor Toll-like receptor 2 (TLR2). Interestingly, the C. rodentium Delta picC mutant caused a degree of pathology similar to that of wild-type C. rodentium when infecting TLR2-deficient mice, showing that despite its mucinase activity, PicC's major role in vivo may be to limit C. rodentium's stimulation of the host's innate immune system.