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Projekt Druckansicht

Insektizide Toxine aus Yersinia enterocolitica

Fachliche Zuordnung Parasitologie und Biologie der Erreger tropischer Infektionskrankheiten
Afrika-, Amerika- und Ozeanienbezogene Wissenschaften
Förderung Förderung von 2008 bis 2021
Projektkennung Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 58161030
 
Erstellungsjahr 2022

Zusammenfassung der Projektergebnisse

The human pathogen Y. enterocolitica strain W22703 exhibits insecticidal and nematocidal activity. This phenotype is caused by proteins of the toxin complex (Tc) encoded by the pathogenicity island Tc-PAIYe. The island genes code for two regulators, TcaR1 and TcaR2, and for the toxin subunits. They are expressed at environmental, but silenced at body temperature. The three subunits form a tripartite ABC-type toxin complex, with subunit C causing cell death through its ADP- ribosyltransferase activity. Molecular and pathophysiological details of insect larvae infection and killing by strain W22703, and the mechanism by which the Tc proteins are translocated across the bacterial membranes lacked disclosure. We first demonstrated biological functionality of the lysis cassette encoded by Tc-PAIYe. Overexpression of the holin/endolysin system in a set of bacterial strains, together with zymography performed with purified endolysin and cell membrane preparations, revealed the specificity of the cassette. The Lon protease was shown to play a role in the control of the temperature-dependent lysis in Y. enterocolitica W22703 at 15°C, possibly by degrading the holin and/or the endolysin. Furthermore, experimental evidence was provided that Tc release requires the lysis cassette, although Y. enterocolitica cell lysis was not observed as a prerequisite for toxin release. We then used larvae of the Greater waxmoth, G. mellonella, to establish an oral model of infection that allowed us to follow the natural route of infection. Tissue sections enabled us to monitor the histopathological impact of the Y. enterocolitica infection, and to visualize the pathogen and its mutants within host compartments by immunofluorescence Time course experiments identified distinct Y. enterocolitica infection stages starting with gut colonisation followed by invasion of the hemolymph, where the pathogen strongly proliferated. The in vivo transcriptome of strain W22703 was delineated after immunomagnetic separation of Y. enterocolitica cells from insect larvae. The transcriptional response was mainly characterized by a drastic reprogramming of the energy, amino acid and carbohydrate metabolism, by an increase of motility and signaling molecules, and by cell membrane rearrangements. Several phenotypes including penetration of the gut epithelium and proliferation in the hemolymph were shown to depend on the presence of the Tc proteins. Strikingly, a mutant lacking a phage-related holin/endolysin (HE) cassette resembled the phenotypes of W22703 ∆tcaA in all readouts of the model, including proliferation and toxicity against the insect, demonstrating that the insecticidal activity conferred by the Tc is directly linked with the lysis cassette. To conclude, the data obtained in this DFG-funded project demonstrate the pivotal role of the Tc proteins in the infection and killing of insect larvae, the specific transcriptional adaptation of Y. enterocolitica to the invertebrate host, and the function of the holin/endolysin system for the release of a high-weight bacterial toxin.

Projektbezogene Publikationen (Auswahl)

 
 

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