Effects of lng Mutations on LngA Expression, Processing, and CS21 Assembly in Enterotoxigenic Escherichia coli E9034A

Abstract

Enterotoxigenic Escherichia coli (ETEC) is a major cause of morbidity in children under 5 years of age in low- and middle-income countries and a leading cause of traveler's diarrhea worldwide. The ability of ETEC to colonize the intestinal epithelium is mediated by fimbrial adhesins, such as CS21 (Longus). This adhesin is a type IVb pilus involved in adherence to intestinal cells in vitro and bacterial self-aggregation. Fourteen open reading frames have been proposed to be involved in CS21 assembly, hitherto only the lngA and lngB genes, coding for the major (LngA) and minor (LngB) structural subunit, have been characterized. In this study, we investigated the role of the LngA, LngB, LngC, LngD, LngH, and LngP proteins in the assembly of CS21 in ETEC strain E9034A. The deletion of the lngA, lngB, lngC, lngD, lngH, or lngP genes, abolished CS21 assembly in ETEC strain E9034A and the adherence to HT-29 cells was reduced 90%, compared to wild-type strain. Subcellular localization prediction of CS21 proteins was similar to other well-known type IV pili homologs. We showed that LngP is the prepilin peptidase of LngA, and that ETEC strain E9034A has another peptidase capable of processing LngA, although with less efficiency. Additionally, we present immuno-electron microscopy images to show that the LngB protein could be localized at the tip of CS21. In conclusion, our results demonstrate that the LngA, LngB, LngC, LngD, LngH, and LngP proteins are essential for CS21 assembly, as well as for bacterial aggregation and adherence to HT-29 cells.

The Vibrio cholerae Minor Pilin TcpB Initiates Assembly and Retraction of the Toxin-Coregulated Pilus

“…The role of the single minor pilin in initiating pilus assembly was shown previously in the CFA/III and Longus T4b pilus systems of ETEC [98, 99, 108], which are closely related to V . cholerae TCP in sequence, structure, gene synteny and pilus functions [98, 109].…”

Interplay of a secreted protein with type IVb pilus for efficient enterotoxigenic Escherichia coli colonization

“…The crystal structure of CofB in complex with a CofJ N-terminal 24-residue synthetic peptide revealed an unprecedented binding mode, by which the peptide is deeply embedded into the expected GalNAc-binding pocket typically conserved at the trimeric interface of the H-type lectin family ( 25 ). This finding provides a conceptual advance in T4P biology, wherein T4bP anchors the secreted protein at the pilus tip for pathogenesis, as well as a plausible answer for the currently debated association state—either monomeric or trimeric—of the type IVb minor pilin in action ( 24 , 32 – 34 ), based on the evidence that the trimeric association of CofB is prerequisite for its stable interaction with CofJ ( 32 – 35 ).…”

“…4 A and B ). The presence of an aromatic patch is commonly observed in PFTs and might act as a multivalent lipid-binding site that recognizes lipid head groups of the cell membrane ( 39 – 41 ), and CofJ reportedly can bind lipid vesicles as well as epithelial cells, such as HeLa and Caco-2 ( 34 ). The lipid-binding nature of CofJ provides advantages for the bacteria by reserving the preferential adhesion spot for initial and/or future attachment to the epithelium as well as in competing with other bacterial species, including commensal species, for subsequent microcolony formation ( Fig.…”

Health Benefits of Lactic Acid Bacteria (LAB) Fermentates

“…Indeed, due to the LAB-associated fermented foods, the gut microbiota can also be altered in a positive manner and such foods can thereby assist in the resolution of IBD [ 30 , 31 ]. It has been suggested that fermented dairy products have the ability to positively modulate the gut microbiota, either by stimulating the proliferation of beneficial microbes and/or by introducing novel species into the gut [ 32 , 33 , 34 ]. Other investigators have studied the hypocholesteremic and anti-cancer properties of these LAB-associated fermented as well as non-fermented milks [ 35 ].…”