Improved gene targeting in Magnaporthe grisea by inactivation of MgKU80 required for non-homologous end joining

• Published in: Fungal genetics and biology Vol. 45; no. 1; pp. 68 - 75
• Main Authors: Villalba, François, Collemare, Jérôme, Landraud, Patricia, Lambou, Karine, Brozek, Viviane, Cirer, Bénédicte, Morin, Damien, Bruel, Christophe, Beffa, Roland, Lebrun, Marc-Henri
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 2008; Elsevier
• Subjects: Ascomycetes; DNA repair; DNA-Binding Proteins - genetics; Fungal Proteins - genetics; Gene Silencing; gene targeting; Gene Targeting - methods; genes; Genes, Fungal; genetic recombination; genetic transformation; Homologous recombination; KU80; Life Sciences; loci; Magnaporthe - genetics; Magnaporthe - growth & development; Magnaporthe - pathogenicity; Magnaporthe grisea; Microbiology and Parasitology; mutagenesis; mutants; Mycology; Neurospora; Non-homologous end joining; nonhomologous end joining; Oryza - microbiology; Phytopathology and phytopharmacy; Plant Diseases - microbiology; Recombination, Genetic; targeted gene replacement; Vegetal Biology; Virulence; KU80; Magnaporthe grisea; Homologous recombination; Non-homologous end joining; MAGNAPORTHE GRISEA; HOMOLOGOUS RECOMBINATION; GENETIQUE; NON-HOMOLOGOUS END JOINING
• ISSN: 1087-1845; 1096-0937
• DOI: 10.1016/j.fgb.2007.06.006

The ascomycete
Magnaporthe grisea is a model species for the study of plant fungal interactions. As in many filamentous fungi, targeted gene replacement occurs at low frequency in
M. grisea (average 7%).
mus52/
KU80 is a gene essential for non-homologous end joining (NHEJ) of DNA double-strand breaks. Its deletion increases the frequency of targeted gene replacement in fungi [Ninomiya, Y., Suzuki, K., Ishii, C., Inoue, H., 2004. Highly efficient gene replacements in
Neurospora strains deficient for non-homologous end joining. Proc. Natl. Acad. Sci. USA 101(33), 12248–53].
M. grisea KU80 deletion mutants were constructed and displayed wild-type phenotypes regarding pathogenicity, growth, sporulation and mating.
MgADE4 targeted gene replacement frequency was increased in Δ
ku80 mutants (80% vs 5%) and high frequencies (>80%) were observed at seven other loci. However, the deletion of
MgKU80 did not increase the frequency of
ACE1 replacement indicating that this locus has an intrinsic reduced ability for gene replacement. These results open the way to large-scale reverse genetics experiments in
M. grisea facilitating the study of the infection process.