TccP4: a Novel Effector Identified in the Strain 1551-2 Required for Attaching and Effacing Lesion Formation on Infected Nck-null Cells

Overview

Journal Microbiol Spectr

Date 2025 Jan 29

PMID 39878470

Authors

Iranildo do A Fernandes

Tadasuke Ooka

Daiany R P de Lira

Fernando H Martins

Henrique Orsi

Nina Jones

Waldir P Elias

Tetsuya Hayashi

Tania A T Gomes

Rodrigo T Hernandes

Affiliations

Soon will be listed here.

Abstract

is a pathogen that causes sporadic cases and outbreaks of diarrhea. The main virulence feature of this bacterium is the attaching and effacing (AE) lesion formation on infected intestinal epithelial cells, which is characterized by the formation of pedestal-like structures that are rich in F-actin. The Brazilian 1551-2 strain can recruit F-actin using both the Nck-dependent and the Nck-independent pathways, the latter of which uses an adaptor protein named Tir-cytoskeleton coupling protein (TccP/EspF). Genome analyses of the 1551-2 strain unveiled the existence of a gene encoding a putative novel TccP subtype in addition to a gene encoding for the TccP3 subtype. Amino-acid sequence comparison with known TccP subtypes (TccP/EspF, TccP2, and TccP3) confirmed that the protein represents a novel TccP subtype-named here TccP4. Lack of TccP4 led to an approximately 96% reduction in the ability of the deletion mutant of strain 1551-2 to induce the F-actin-rich pedestals formation in the infected Nck-null mouse embryonic fibroblasts (MEF) cells. The gene was distributed widely in , including the strains first separated from other strains, suggesting that this gene was acquired at a very early stage during the diversification of . The highly variable genetic organization of the -containing regions and the presence of various mobile genetic elements in this region may explain the lack of in strains belonging to various lineages.IMPORTANCE, one of the new members of the genus is a diarrheagenic pathogen. The main characteristic of its pathogenicity is the formation of attaching and effacing (AE) lesions on the surface of infected epithelial cells. Here we identified a novel subtype of the TccP type 3 secretion system (T3SS) effector family (termed TccP4), which is required for the recruitment of F-actin during the AE lesion formation in infected host cells by the 1551-2 strain. We also revealed that TccP4 is unique to and widely distributed in this species, suggesting that the gene was acquired at a very early stage during the diversification process of . These findings expand our understanding of the function and diversity of this important T3SS effector family.

References

Stamatakis A . RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics. 2014; 30(9):1312-3. PMC: 3998144. DOI: 10.1093/bioinformatics/btu033. View

Moon H, Whipp S, Argenzio R, Levine M, Giannella R . Attaching and effacing activities of rabbit and human enteropathogenic Escherichia coli in pig and rabbit intestines. Infect Immun. 1983; 41(3):1340-51. PMC: 264644. DOI: 10.1128/iai.41.3.1340-1351.1983. View

Datsenko K, Wanner B . One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A. 2000; 97(12):6640-5. PMC: 18686. DOI: 10.1073/pnas.120163297. View

Garmendia J, Phillips A, Carlier M, Chong Y, Schuller S, Marches O . TccP is an enterohaemorrhagic Escherichia coli O157:H7 type III effector protein that couples Tir to the actin-cytoskeleton. Cell Microbiol. 2004; 6(12):1167-83. DOI: 10.1111/j.1462-5822.2004.00459.x. View

Kenny B . Phosphorylation of tyrosine 474 of the enteropathogenic Escherichia coli (EPEC) Tir receptor molecule is essential for actin nucleating activity and is preceded by additional host modifications. Mol Microbiol. 1999; 31(4):1229-41. DOI: 10.1046/j.1365-2958.1999.01265.x. View

Parks D, Imelfort M, Skennerton C, Hugenholtz P, Tyson G . CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res. 2015; 25(7):1043-55. PMC: 4484387. DOI: 10.1101/gr.186072.114. View

Crepin V, Girard F, Schuller S, Phillips A, Mousnier A, Frankel G . Dissecting the role of the Tir:Nck and Tir:IRTKS/IRSp53 signalling pathways in vivo. Mol Microbiol. 2009; 75(2):308-23. PMC: 2814079. DOI: 10.1111/j.1365-2958.2009.06938.x. View

Yamamoto D, Hernandes R, Liberatore A, Abe C, Souza R, Romao F . Escherichia albertii, a novel human enteropathogen, colonizes rat enterocytes and translocates to extra-intestinal sites. PLoS One. 2017; 12(2):e0171385. PMC: 5298312. DOI: 10.1371/journal.pone.0171385. View

Jones D, Taylor W, Thornton J . The rapid generation of mutation data matrices from protein sequences. Comput Appl Biosci. 1992; 8(3):275-82. DOI: 10.1093/bioinformatics/8.3.275. View

10.

Letunic I, Bork P . Interactive tree of life (iTOL) v3: an online tool for the display and annotation of phylogenetic and other trees. Nucleic Acids Res. 2016; 44(W1):W242-5. PMC: 4987883. DOI: 10.1093/nar/gkw290. View

11.

Kenny B, DeVinney R, Stein M, Reinscheid D, Frey E, Finlay B . Enteropathogenic E. coli (EPEC) transfers its receptor for intimate adherence into mammalian cells. Cell. 1997; 91(4):511-20. DOI: 10.1016/s0092-8674(00)80437-7. View

12.

Laemmli U . Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970; 227(5259):680-5. DOI: 10.1038/227680a0. View

13.

Gaytan M, Martinez-Santos V, Soto E, Gonzalez-Pedrajo B . Type Three Secretion System in Attaching and Effacing Pathogens. Front Cell Infect Microbiol. 2016; 6:129. PMC: 5073101. DOI: 10.3389/fcimb.2016.00129. View

14.

Scita G, Confalonieri S, Lappalainen P, Suetsugu S . IRSp53: crossing the road of membrane and actin dynamics in the formation of membrane protrusions. Trends Cell Biol. 2008; 18(2):52-60. DOI: 10.1016/j.tcb.2007.12.002. View

15.

Aitio O, Hellman M, Kazlauskas A, Vingadassalom D, Leong J, Saksela K . Recognition of tandem PxxP motifs as a unique Src homology 3-binding mode triggers pathogen-driven actin assembly. Proc Natl Acad Sci U S A. 2010; 107(50):21743-8. PMC: 3003032. DOI: 10.1073/pnas.1010243107. View

16.

Chaveroche M, Ghigo J, dEnfert C . A rapid method for efficient gene replacement in the filamentous fungus Aspergillus nidulans. Nucleic Acids Res. 2000; 28(22):E97. PMC: 113889. DOI: 10.1093/nar/28.22.e97. View

17.

Hernandes R, Silva R, Carneiro S, Salvador F, Fernandes M, Padovan A . The localized adherence pattern of an atypical enteropathogenic Escherichia coli is mediated by intimin omicron and unexpectedly promotes HeLa cell invasion. Cell Microbiol. 2007; 10(2):415-25. DOI: 10.1111/j.1462-5822.2007.01054.x. View

18.

Lima M, Yamamoto D, Santos A, Ooka T, Hernandes R, Vieira M . Phenotypic characterization and virulence-related properties of Escherichia albertii strains isolated from children with diarrhea in Brazil. Pathog Dis. 2019; 77(2). DOI: 10.1093/femspd/ftz014. View

19.

Cheng H, Skehan B, Campellone K, Leong J, Rosen M . Structural mechanism of WASP activation by the enterohaemorrhagic E. coli effector EspF(U). Nature. 2008; 454(7207):1009-13. PMC: 2719906. DOI: 10.1038/nature07160. View

20.

Knutton S, Baldwin T, Williams P, McNeish A . Actin accumulation at sites of bacterial adhesion to tissue culture cells: basis of a new diagnostic test for enteropathogenic and enterohemorrhagic Escherichia coli. Infect Immun. 1989; 57(4):1290-8. PMC: 313264. DOI: 10.1128/iai.57.4.1290-1298.1989. View