CRISPRi in Xanthomonas demonstrates functional convergence of transcription activator-like effectors in two divergent pathogens

Zárate-Chaves Carlos Andrés, Audran Corinne, Medina Culma César Augusto, Escalon Aline, Javegny Stéphanie, Gagnevin Lionel, Thomas Emilie, Pimparé Léa-Lou, López Camilo, Jacobs Jonathan M., Noel Laurent D., Koebnik Ralf, Bernal Adriana Jimena, Szurek Boris. 2023. CRISPRi in Xanthomonas demonstrates functional convergence of transcription activator-like effectors in two divergent pathogens. New Phytologist, 238 (4) : 1593-1604.

https://doi.org/10.1111/nph.18808

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Url - jeu de données - Entrepôt autre : https://www.ncbi.nlm.nih.gov/datasets/genome/GCF_000019585.2/ / Url - jeu de données - Entrepôt autre : https://www.ncbi.nlm.nih.gov/datasets/genome/GCF_003031365.1/ / Url - jeu de données - Entrepôt autre : https://www.ncbi.nlm.nih.gov/datasets/genome/GCF_000007165.1/

Résumé : Functional analysis of large gene families in plant pathogens can be cumbersome using classical insertional mutagenesis. Additionally, Cas9 toxicity has limited the application of CRISPR–Cas9 for directed mutagenesis in bacteria. Here, we successfully applied a CRISPR interference strategy to investigate the cryptic role of the transcription activator-like effector (tale) multigene family in several plant-pathogenic Xanthomonas bacterial species, owing to their contribution to pathogen virulence. Single guide RNAs (sgRNAs) designed against Xanthomonas phaseoli pv manihotis tale conserved gene sequences efficiently silenced expression of all tales, with concomitant decrease in virulence and TALE-induced host gene expression. The system is readily translatable to other Xanthomonas species infecting rice, citrus, Brassica, and cassava, silencing up to 16 tales in a given strain using a single sgRNA. Complementation with plasmid-borne designer tales lacking the sgRNA-targeted sequence restored molecular and virulence phenotypes in all pathosystems. Our results evidenced that X. campestris pv campestris CN08 tales are relevant for symptom development in cauliflower. They also show that the MeSWEET10a sugar transporter is surprisingly targeted by the nonvascular cassava pathogen X. cassavae, highlighting a new example of TALE functional convergence between phylogenetically distant Xanthomonas. Overall, this novel technology provides a platform for discovery and rapid functional understanding of highly conserved gene families.

Mots-clés Agrovoc : Oryza sativa, Xanthomonas, expression des gènes, phylogénie, Xanthomonas oryzae, Transcription génique, virulence, Manihot esculenta, pouvoir pathogène, Xanthomonas campestris manihotis, Xanthomonas campestris phaseoli, effecteur moléculaire, maladie des plantes, gène, phénotype