Listeria monocytogenes causes invasive disease by crossing the intestinal epithelial barrier. This process depends on the interaction between the bacterial surface protein Internalin A (InlA) and the host protein E-cadherin. A second L. monocytogenes invasin Internalin B (InlB) promotes invasion of numerous non-phagocytic cell types, but has not been shown to promote oral infection. The receptor for InlB is c-Met, a receptor tyrosine kinase and the endogenous receptor for Hepatocyte Growth Factor (HGF). E-cadherin and c-Met are localized to the basolateral side of polarized epithelial cells and are not thought to be accessible to the apical (lumenal) side across functional tight junctions.
We used polarized MDCK cells as a model epithelium to determine how L. monocytogenes gain access to basolateral receptors. We found that L. monocytogenes do not actively disrupt the tight junctions, but find E-cadherin at a morphologically distinct subset of intercellular junctions. We identified these sites as naturally occurring regions where single senescent cells are extruded from the epithelium. The surrounding cells reorganize to form a multicellular junction (MCJ) that maintains epithelial continuity. We found that E-cadherin is transiently exposed to the lumenal surface at MCJs during and after cell extrusion.
We hypothesized that L. monocytogenes utilize analogous extrusion sites for epithelial invasion in vivo. By infecting rabbit ileal loops, we found that the MCJs at the cell extrusion zone of villus tips are the specific target for InlA-mediated L. monocytogenes adhesion and invasion. L. monocytogenes expressing a modified InlA capable of binding murine E-cadherin (InlAm) specifically invade and replicate within villous tips of orally infected of mice. We hypothesized that InlB functions synergistically with Inla to promote intestinal invasion. Utilizing L. monocytogenes expressing InlAm, we found that InllB promotes oral infection of mice and colonization of mouse villous tips.
We investigated the mechanism by which InlB mediates Listeria invasion at MCJs using polarized MDCK monolayers. Following InlA-mediated attachment at MCJs, c-Met activation by InlB increases the rate of bacterial uptake. The efficiency of invasion is also controlled by intrinsic epithelial properties since MCJs undergo rapid remodeling and are naturally more endocytic than other junctional sites; MCJs endocytose fluorescent dextran, a fluid phase marker, from the apical surface into unique cytoplasmic puncta containing both tight- and adherens junction proteins. Apical HGF or InlB increase the number and size of dextran puncta at MCJs, but do not increase endocytosis at other junctions, suggesting that c-Met is apically exposed at MCJs and that L. monocytogenes can modulate cellular endocytosis during invasion of this specific site.
Thus, L. monocytogenes exploit the dynamic nature of junctional remodeling and epithelial renewal to target exposed receptors and hijack host cell processes for epithelial invasion and intestinal barrier breach.
|School Location:||United States -- California|
|Source:||DAI-B 70/10, Dissertation Abstracts International|
|Subjects:||Molecular biology, Microbiology, Immunology|
|Keywords:||E-cadherin, Endocytosis, Intestinal epithelium, Listeria|
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