Enteropathogenic Escherichia coli dephosphorylates and dissociates occludin from intestinal epithelial tight junctions

Enteropathogenic Escherichia coli (EPEC) increases tight junction permeabil ity in part by phosphorylating the 20 kDa myosin light chain (MLC20) that i nduces cytoskeletal contraction. The impact of this enteric pathogen on spe cific tight junction (TJ) proteins has not been investigated. We examined t he effect of EPEC infection on occludin localization and phosphorylation in intestinal epithelial cells. After infection by EPEC, a progressive shift of occludin from a primarily TJ-associated domain to an intracellular compa rtment occurred, as demonstrated by immunofluorescent staining. A reverse i n the ratio of phosphorylated to dephosphorylated occludin accompanied this morphological change. Eradication of EPEC with gentamicin resulted in the normalization of occludin localization and phosphorylation. The serine/thre onine phosphatase inhibitor, calyculin A, prevented these events. The EPEC- associated decrease in transepithelial electrical resistance, a measure of TJ barrier function, returned to baseline after gentamicin treatment. Non-p athogenic E. coli, K-12, did not induce these changes. Transformation of K- 12 with the pathogenicity island of EPEC, however, conferred the phenotype of wild-type EPEC. Deletion of specific EPEC genes encoding proteins involv ed in EPEC type III secretion markedly attenuated these effects. These find ings suggest that EPEC-induced alterations in occludin contribute to the pa thophysiology associated with this infection.