ARS | Publication request: The EHEC type III effector NleL is an E3 ubiquitin ligase that modulates pedestal formation

Submitted to: PLoS One
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 25, 2011
Publication Date: April 26, 2011
Repository URL: http://www.ncbi.nlm.nih.gov/pubmed/21541301
Citation: Piscatelli, H., Kotkar, S.A., Mcbee, M.E., Muthupalani, S., Schauer, D.B., Mandrell, R.E., Leong, J.M., Chen, J., Zhou, D. 2011. The EHEC type III effector NleL is an E3 ubiquitin ligase that modulates pedestal formation. PLoS One. 6:e19331.

Interpretive Summary: Enterohemorrhagic Escherichia coli (EHEC) O157:H7 causes hemorrhagic colitis and may result in potentially fatal hemolytic uremia syndrome in humans. Similar to members of the attaching and effacing (A/E) pathogens, EHEC colonize the intestinal mucosa and promote the formation of actin-rich pedestals beneath the adherent bacteria via the trasnlocated type III effectors. Two EHEC type III secreted effectors, Tir and EspFu/TccP, are known to be the key players for the pedestal formation. Bacterial encoded intimin binds to Tir on the plasma membrane of the host cells to promote the formation of actin-rich pedestals within the host cell beneath the adherent bacteria. We discovered that an effector protein called Non-LEE-encoded Ligase (NleL) encoded by the ECs1560 locus of EHEC is an E3 ubiquitin ligase. In vitro, we showed that the NleL C753 residue is critical for its E3 ligase activity. Functionally, we demonstrated that NleL and its E3 ubiquitin ligase activity are involved in modulating Tir-mediated pedestal formation. Surprisingly, EHEC mutant strain deficient in the E3 ligase activity induced more pedestals when compared to that induced by the wild-type strain. EPEC strain, which lacks the nleL gene, induced less actin-rich pedestals when carrying the wild-type EHEC nleL compared to EPEC carrying the catalytically deficient nleL (C753S) mutant. Furthermore, we showed that NleLc, a homolog of NleL from Citrobacter rodentium, is a bona fide E3 ubiquitin ligase and that the E3 ligase activity of NleLc is required for efficient infection of murine colonic epithelial cells in vivo. In summary, our study demonstrated that EHEC utilizes NleL E3 ubiquitin ligase activity to modulate the efficiency of Tir-mediated pedestal formation.

Technical Abstract: Enterohemorrhagic Escherichia coli (EHEC) O157:H7 causes hemorrhagic colitis and may result in potentially fatal hemolytic uremia syndrome in humans. EHEC colonize the intestinal mucosa and promote formation of “pedestals” in the tissue beneath the adherent bacteria. Secreted proteins are key players for the pedestal formation. We discovered that a specific effector protein called Non-LEE-encoded Ligase (NleL) is an ubiquitin ligase involved in modulating Tir-mediated pedestal formation. A homolog of NleL from Citrobacter rodentium also has E3 ligase activity and is required for efficient infection of murine colonic epithelial cells in vivo. In summary, our study demonstrated that EHEC utilizes NleL E3 ubiquitin ligase activity to modulate the efficiency of Tir-mediated pedestal formation.