The akuB ^KU80 Mutant Deficient for Nonhomologous End Joining Is a Powerful Tool for Analyzing Pathogenicity in Aspergillus fumigatus

Abstract

To increase the frequency of homologous recombination, we inactivated the KU80 homologue in Aspergillus fumigatus (named akuB KU80 ). Homologous integration reached about 80% for both calcineurin A (calA) and polyketide synthase pksP (alb1) genes in the akuB KU80 mutant to 3 and 5%, respectively, when using a wild-type A. fumigatus strain. Deletion of akuB KU80 had no influence on pathogenicity in a low-dose murine infection model. Double-strand breaks are the most severe form of DNA damage. Eukaryotes have two main pathways to deal with this type of DNA damage: (i) homologous recombination, involving interaction between homologous sequences, and (ii) nonhomologous end joining, involving direct ligation of the strand ends independently of DNA homology (17). The most important double-strand break repair pathway in Saccharomyces cerevisiae is homologous recombination (18,20), while other organisms, such as humans, preferentially use nonhomologous end joining. The nonhomologous end-joining process is mediated by the DNA-dependent protein kinase catalytic subunit, the Ku70-Ku80 heterodimer, and the DNA ligase IV-Xrcc4 complex (17). Recently, Ninomiya et al. (13) disrupted Neurospora crassa genes homologous to human KU70 and KU80.

An alternative host model of a mixed fungal infection by azole susceptible and resistantAspergillusspp strains

“…fumigatus (akuB KU80 ), 17 CM-237, 18 AF14 16 and A. lentulus (CM-1290) strains were selected for our infection model and further analysis.…”

Contributions of Aspergillus fumigatus ATP-Binding Cassette Transporter Proteins to Drug Resistance and Virulence

“…fumigatus strains, growth conditions, and transformation. Three strains were used in this study: the A. fumigatus Af293 strain, for which the entire genomic sequence is available (23); the akuB⌬ (A1151) strain (24); and the akuA⌬ (AfS35(strain (25)). These strains were typically grown at 37°C in rich medium (YG; 0.5% yeast extract, 2% glucose) or in minimal medium (MM; 1% glucose, nitrate salts, trace elements, 2% agar [pH 6.5]; trace elements, vitamins, and nitrate salts are as described in the appendix of reference 26.…”

The putative flavin carrier family FlcA-C is important forAspergillus fumigatusvirulence

“…52,53 All the comparisons with the deletion strains were performed with the CEA17 pyrG C . The media used were: complete medium composed for 2% w/v glucose, 0.5% w/v yeast extract, 2% w/v agar, trace elements (YAG) or YUU [YAG supplemented with 1.2 g (each) of uracil and uridine], and liquid YG or YG C UU medium with the same composition (but without agar).…”