Hfq and three Hfq-dependent small regulatory RNAs—MgrR, RyhB and McaS—coregulate the locus of enterocyte effacement in enteropathogenicEscherichia coli

Abstract

Enteropathogenic Escherichia coli (EPEC) is a significant cause of infantile diarrhea and death in developing countries. The pathogenicity island locus of enterocyte effacement (LEE) is essential for EPEC to cause diarrhea. Besides EPEC, the LEE is also present in other gastrointestinal pathogens, most notably enterohemorrhagic E. coli (EHEC). Whereas transcriptional control of the LEE has been meticulously examined, posttranscriptional regulation, including the role of Hfq-dependent small RNAs, remains undercharacterized. However, the past few years have witnessed a surge in the identification of riboregulators of the LEE in EHEC. Contrastingly, the posttranscriptional regulatory landscape of EPEC remains cryptic. Here we demonstrate that the RNA-chaperone Hfq represses the LEE of EPEC by targeting the 5' untranslated leader region of grlR in the grlRA mRNA. Three conserved small regulatory RNAs (sRNAs)-MgrR, RyhB and McaS-are involved in the Hfq-dependent regulation of grlRA MgrR and RyhB exert their effects by directly base-pairing to the 5' region of grlR Whereas MgrR selectively represses grlR but activates grlA, RyhB represses gene expression from the entire grlRA transcript. Meanwhile, McaS appears to target the grlRA mRNA indirectly. Thus, our results provide the first definitive evidence that implicates multiple sRNAs in regulating the LEE and the resulting virulence of EPEC.

The Small Regulatory RNA Spot42 Inhibits Indole Biosynthesis to Negatively Regulate the Locus of Enterocyte Effacement of Enteropathogenic Escherichia coli

“…Beta-galactosidase assays on the translational fusion ( P araBAD -tnaA’-‘lacZ ) containing reporter strains were performed as described in our previous paper [ 19 ], with the exception that bacteria were grown to an optical density of 1.1–1.4 in LB supplemented with ampicillin, arabinose, and IPTG. The same conditions, as described above, were employed for the transcriptional fusion ( P LEE1 -LEE1’-lacZ + ) containing reporter strains with the omission of arabinose from the medium.…”

“…The T3SS bridges the bacterium to the host cell and facilitates the smuggling of effector proteins into the host where they subvert host signaling pathways to initiate A/E lesion formation [ 4 , 13 , 18 ]. The importance of this genomic island to the bacterium is illustrated by the characterization of over 40 regulatory factors that control the LEE [ 4 , 19 ]. Most regulators target one of the two key regulatory operons— grlRA ( LEE7 ) and/or LEE1.…”

“…In a previous study, we reported the first, and thus far, only sRNAs—MgrR and RyhB—that directly regulate the LEE in EPEC [ 19 ]. The present study was undertaken to identify sRNAs that indirectly regulate the LEE by modulating gene expression from the well-known regulatory operon tnaCAB .…”

After the Fact(or): Posttranscriptional Gene Regulation in Enterohemorrhagic Escherichia coli O157:H7

“…Although the authors extensively characterized the involvement of Hfq in expression of LEE4 (detailed below), no data describing the specific mechanism of Hfq-dependent regulation of Ler or GrlA expression were reported in that study. However, the Hfqdependent sRNAs MgrR and RyhB directly influence LEE expression in enteropathogenic E. coli (112). These sRNAs were identified following a screen that assessed the impact of core sRNAs on the activity of grlR=-=lacZ reporter fusions.…”

“…These sRNAs were identified following a screen that assessed the impact of core sRNAs on the activity of grlR=-=lacZ reporter fusions. Further characterization demonstrated that MgrR base pairs to the 5= UTR of grlR to repress grlR expression and to activate expression of grlA and the LEE, whereas RyhB binds to the grlRA mRNA to repress translation (112) (Fig. 6B).…”