title: Rapid paracellular transmigration of Campylobacter jejuni across polarized epithelial cells without affecting TER: role of proteolytic-active HtrA cleaving E-cadherin but not fibronectin.
abstract: BACKGROUND
  Campylobacter jejuni is one of the most important bacterial pathogens causing food-borne illness worldwide. Crossing the intestinal epithelial barrier and host cell entry by C. jejuni is considered the primary reason of damage to the intestinal tissue, but the molecular mechanisms as well as major bacterial and host cell factors involved in this process are still widely unclear.
  
  
  RESULTS
  In the present study, we characterized the serine protease HtrA (high-temperature requirement A) of C. jejuni as a secreted virulence factor with important proteolytic functions. Infection studies and in vitro cleavage assays showed that C. jejuni's HtrA triggers shedding of the extracellular E-cadherin NTF domain (90 kDa) of non-polarised INT-407 and polarized MKN-28 epithelial cells, but fibronectin was not cleaved as seen for H. pylori's HtrA. Deletion of the htrA gene in C. jejuni or expression of a protease-deficient S197A point mutant did not lead to loss of flagella or reduced bacterial motility, but led to severe defects in E-cadherin cleavage and transmigration of the bacteria across polarized MKN-28 cell layers. Unlike other highly invasive pathogens, transmigration across polarized cells by wild-type C. jejuni is highly efficient and is achieved within a few minutes of infection. Interestingly, E-cadherin cleavage by C. jejuni occurs in a limited fashion and transmigration required the intact flagella as well as HtrA protease activity, but does not reduce transepithelial electrical resistance (TER) as seen with Salmonella, Shigella, Listeria or Neisseria.
  
  
  CONCLUSIONS
  These results suggest that HtrA-mediated E-cadherin cleavage is involved in rapid crossing of the epithelial barrier by C. jejuni via a very specific mechanism using the paracellular route to reach basolateral surfaces, but does not cleave the fibronectin receptor which is necessary for cell entry.
authors.affiliation: School for Medicine and Medical Science, University College Dublin, Belfield Campus, Dublin-4, Ireland.
authors.forename: Manja
authors.initials: M
authors.lastname: Boehm
authors.affiliation: The Division of Microbiology, University Salzburg, A-5020, Salzburg, Austria.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Hoy
authors.affiliation: The Department of Medical Microbiology, Helmholtz Center for Infection Research, Inhoffen Str. 7, D-38124, Braunschweig, Germany.
authors.forename: Manfred
authors.initials: M
authors.lastname: Rohde
authors.affiliation: School for Medicine and Medical Science, University College Dublin, Belfield Campus, Dublin-4, Ireland.
authors.forename: Nicole
authors.initials: N
authors.lastname: Tegtmeyer
authors.affiliation: The Department of Veterinary Disease Biology, University Copenhagen, Stigbøjlen 4, DK-1870, Frederiksberg C, Denmark.
authors.forename: Kristoffer T
authors.initials: KT
authors.lastname: Bæk
authors.affiliation: Institute for Environmental Health, Inc., 15300 Bothell Way NE Lake Forest Park, Seattle, WA, 98155, USA.
authors.forename: Omar A
authors.initials: OA
authors.lastname: Oyarzabal
authors.affiliation: The Department of Veterinary Disease Biology, University Copenhagen, Stigbøjlen 4, DK-1870, Frederiksberg C, Denmark.
authors.forename: Lone
authors.initials: L
authors.lastname: Brøndsted
authors.affiliation: The Division of Microbiology, University Salzburg, A-5020, Salzburg, Austria.
authors.forename: Silja
authors.initials: S
authors.lastname: Wessler
authors.affiliation: School for Medicine and Medical Science, University College Dublin, Belfield Campus, Dublin-4, Ireland.|University College Dublin, UCD School of Biomolecular and Biomedical Sciences, Science Center West L231, Belfield Campus, Dublin 4, Ireland.
authors.forename: Steffen
authors.initials: S
authors.lastname: Backert