Functional differences and interactions between the Escherichia coli type III secretion system effectors NleH1 and...

Area of Science:

• Microbiology
• Immunology
• Molecular Biology

Background:

• Enterohemorrhagic Escherichia coli (EHEC), enteropathogenic E. coli (EPEC), and Citrobacter rodentium employ type III secretion systems (T3SS) to deliver effector proteins into host cells.
• The E. coli O157:H7 strain EDL933 encodes two non-locus of enterocyte effacement (LEE)-encoded proteins, NleH1 and NleH2, which interact with ribosomal protein S3 (RPS3), a component of NF-κB transcriptional complexes.

Purpose of the Study:

• To investigate the functional differences between NleH1 and NleH2 in regulating the host NF-κB pathway.
• To elucidate the role of the PDZ-binding domain in NleH activity.
• To assess the in vivo virulence contributions of NleH1 and NleH2 in a mouse infection model.

Main Methods:

• Assessed the impact of NleH1 and NleH2 on RPS3 nuclear translocation and NF-κB activity.
• Investigated the interaction between NleH1 and NleH2 using in vitro and in vivo assays.
• Performed mouse infection experiments with wild-type, mutant, and complemented strains to evaluate virulence and colonization.

Main Results:

• NleH1, but not NleH2, inhibited NF-κB activity without affecting IκBα phosphorylation/degradation kinetics.
• The class I PDZ-binding domain of NleH was crucial for its NF-κB pathway activity.
• NleH1 and NleH2 were found to bind each other, suggesting a regulatory mechanism for NF-κB activation.
• Mice infected with a ΔnleH mutant showed reduced mortality and Citrobacter colonization; NleH1 complemented virulence, while NleH2 did not.

Conclusions:

• NleH1 and NleH2 possess distinct functional roles in modulating host NF-κB signaling.
• The interaction between NleH1 and NleH2 may fine-tune NF-κB activation.
• NleH1 is the primary driver of Citrobacter rodentium virulence in the studied mouse model, while NleH2 plays a lesser role.