Analysis of the expression, regulation and export of NleA-E in Escherichia coli O157: H7

Previous work has shown that locus of enterocyte effacement (LEE)-encoded effector proteins such as Tir and Map can be exported via the type III secretion system (T3SS) of Escherichia coli O157 : H7. Additionally, a family of non-LEE-encoded (Nle) effector proteins has been shown to be secreted from Citrobacter rodentium, homologues of which are located on the E. coli O157 chromosome. While NleA has been shown to be secreted from pathogenic E. coli, the secretion of other Nle effector proteins has only been detected under induced conditions, or using a mutated T3SS. This study aimed to determine: (1) which nle genes are expressed in E. coli O157 : H7 under secretion-permissive conditions; (2) if Nle proteins are secreted from wild-type E. coli O157 : H7 under secretion-permissive conditions; and (3) if nle gene expression is regulated co-ordinately with other LEE-encoded effectors. Using data generated from a combination of transcriptome arrays, reporter fusions and proteomics, it was demonstrated that only nleA is expressed co-ordinately with the LEE. Secretion and expression of NleA were regulated directly or indirectly by ler, a key activator of the LEE. MS confirmed the secretion of NleA into the culture supernatant, while NleB-F were not detected.