ABSTRACT
Genetic stability is extremely important for the survival of every living organism, and a very complex set of genes has evolved to cope with DNA repair upon DNA damage. Here, we investigated the Aspergillus fumigatus AtmA (Ataxia-telangiectasia mutated, ATM) and AtrA kinases, and how they impact virulence and the evolution of azole resistance. We demonstrated that A. fumigatus atmA and atrA null mutants are haploid and have a discrete chromosomal polymorphism. The ΔatmA and ΔatrA strains are sensitive to several DNA-damaging agents, but surprisingly both strains were more resistant than the wild-type strain to paraquat, menadione, and hydrogen peroxide. The atmA and atrA genes showed synthetic lethality emphasizing the cooperation between both enzymes and their consequent redundancy. The lack of atmA and atrA does not cause any significant virulence reduction in A. fumigatus in a neutropenic murine model of invasive pulmonary aspergillosis and in the invertebrate alternative model Galleria mellonela Wild-type, ΔatmA, and ΔatrA populations that were previously transferred 10 times in minimal medium (MM) in the absence of voriconazole have not shown any significant changes in drug resistance acquisition. In contrast, ΔatmA and ΔatrA populations that similarly evolved in the presence of a subinhibitory concentration of voriconazole showed an â¼5-10-fold increase when compared to the original minimal inhibitory concentration (MIC) values. There are discrete alterations in the voriconazole target Cyp51A/Erg11A or cyp51/erg11 and/or Cdr1B efflux transporter overexpression that do not seem to be the main mechanisms to explain voriconazole resistance in these evolved populations. Taken together, these results suggest that genetic instability caused by ΔatmA and ΔatrA mutations can confer an adaptive advantage, mainly in the intensity of voriconazole resistance acquisition.
Subject(s)
Aspergillus fumigatus/drug effects , Ataxia Telangiectasia Mutated Proteins/genetics , Drug Resistance, Fungal/genetics , Gene Expression Regulation, Fungal , Genome, Fungal , Voriconazole/pharmacology , Animals , Antifungal Agents/pharmacology , Aspergillus fumigatus/genetics , Aspergillus fumigatus/metabolism , Ataxia Telangiectasia Mutated Proteins/deficiency , Cytochrome P-450 Enzyme System/genetics , Cytochrome P-450 Enzyme System/metabolism , Disease Models, Animal , Female , Fungal Proteins/genetics , Fungal Proteins/metabolism , Genomic Instability , Humans , Invasive Pulmonary Aspergillosis/drug therapy , Invasive Pulmonary Aspergillosis/microbiology , Invasive Pulmonary Aspergillosis/mortality , Invasive Pulmonary Aspergillosis/pathology , Membrane Transport Proteins/genetics , Membrane Transport Proteins/metabolism , Mice , Mice, Inbred BALB C , Microbial Sensitivity Tests , Moths/microbiology , Mutation , Protein Isoforms/deficiency , Protein Isoforms/genetics , Survival Analysis , VirulenceABSTRACT
Nibrin (NBN or NBS1) and ATM are key factors for DNA Double Strand Break (DSB) signaling and repair. Mutations in NBN or ATM result in Nijmegen Breakage Syndrome and Ataxia telangiectasia. These syndromes share common features such as radiosensitivity, neurological developmental defects and cancer predisposition. However, the functional synergy of Nbn and Atm in different tissues and developmental stages is not yet understood. Here, we show in vivo consequences of conditional inactivation of both genes in neural stem/progenitor cells using Nestin-Cre mice. Genetic inactivation of Atm in the central nervous system of Nbn-deficient mice led to reduced life span and increased DSBs, resulting in increased apoptosis during neural development. Surprisingly, the increase of DSBs and apoptosis was found only in few tissues including cerebellum, ganglionic eminences and lens. In sharp contrast, we showed that apoptosis associated with Nbn deletion was prevented by simultaneous inactivation of Atm in developing retina. Therefore, we propose that Nbn and Atm collaborate to prevent DSB accumulation and apoptosis during development in a tissue- and developmental stage-specific manner.