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Molecular characterization of an adaptive response to alkylating agents in the opportunistic pathogen Aspergillus fumigatus.

ABSTRACT: An adaptive response to alkylating agents based upon the conformational change of a methylphosphotriester (MPT) DNA repair protein to a transcriptional activator has been demonstrated in a number of bacterial species, but this mechanism appears largely absent from eukaryotes. Here, we demonstrate that the human pathogen Aspergillus fumigatus elicits an adaptive response to sub-lethal doses of the mono-functional alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). We have identified genes that encode MPT and O(6)-alkylguanine DNA alkyltransferase (AGT) DNA repair proteins; deletions of either of these genes abolish the adaptive response and sensitize the organism to MNNG. In vitro DNA repair assays confirm the ability of MPT and AGT to repair methylphosphotriester and O(6)-methylguanine lesions respectively. In eukaryotes, the MPT protein is confined to a select group of fungal species, some of which are major mammalian and plant pathogens. The evolutionary origin of the adaptive response is bacterial and rooted within the Firmicutes phylum. Inter-kingdom horizontal gene transfer between Firmicutes and Ascomycete ancestors introduced the adaptive response into the Fungal kingdom. Our data constitute the first detailed characterization of the molecular mechanism of the adaptive response in a lower eukaryote and has applications for development of novel fungal therapeutics targeting this DNA repair system.

SUBMITTER: O'Hanlon KA

PROVIDER: S-EPMC3439912 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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Molecular characterization of an adaptive response to alkylating agents in the opportunistic pathogen Aspergillus fumigatus.

O'Hanlon Karen A KA Margison Geoffrey P GP Hatch Amy A Fitzpatrick David A DA Owens Rebecca A RA Doyle Sean S Jones Gary W GW

Nucleic acids research 20120604 16

An adaptive response to alkylating agents based upon the conformational change of a methylphosphotriester (MPT) DNA repair protein to a transcriptional activator has been demonstrated in a number of bacterial species, but this mechanism appears largely absent from eukaryotes. Here, we demonstrate that the human pathogen Aspergillus fumigatus elicits an adaptive response to sub-lethal doses of the mono-functional alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). We have identified gen ...[more]

PMID: 22669901

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Project description:Aspergillus fumigatus is a potentially deadly opportunistic fungal pathogen. Molecular studies have shaped our understanding of the genes, proteins, and molecules that contribute to A. fumigatus pathogenicity, but few studies have characterized genome-wide patterns of genetic variation at the population level. Of A. fumigatus genomic studies to-date, most focus mainly on single nucleotide polymorphisms and large structural variants, while overlooking the contribution of copy number variation (CNV). CNV is a class of small structural variation defined as loci that vary in their number of copies between individuals due to duplication, gain, or deletion. CNV can influence phenotype, including fungal virulence. In the present study, we characterized the population genomic patterns of CNV in a diverse collection of 71 A. fumigatus isolates using publicly available sequencing data. We used genome-wide single nucleotide polymorphisms to infer the population structure of these isolates and identified three populations consisting of at least 8 isolates. We then computationally predicted genome-wide CNV profiles for each isolate and conducted analyses at the species-, population-, and individual levels. Our results suggest that CNV contributes to genetic variation in A. fumigatus, with ~10% of the genome being CN variable. Our analysis indicates that CNV is non-randomly distributed across the A. fumigatus genome, and is overrepresented in subtelomeric regions. Analysis of gene ontology categories in genes that overlapped CN variants revealed an enrichment of genes related to transposable element and secondary metabolism functions. We further identified 72 loci containing 33 genes that showed divergent copy number profiles between the three A. fumigatus populations. Many of these genes encode proteins that interact with the cell surface or are involved in pathogenicity. Our results suggest that CNV is an important source of genetic variation that could account for some of the phenotypic differences between A. fumigatus populations and isolates.

Project description:Aspergillus nidulans is an opportunistic fungal pathogen in patients with immunodeficiency, and virulence of A. nidulans isolates has mainly been studied in the context of chronic granulomatous disease (CGD), with characterization of clinical isolates obtained from non-CGD patients remaining elusive. This study therefore carried out a detailed biological characterization of two A. nidulans clinical isolates (CIs), obtained from a patient with breast carcinoma and pneumonia and from a patient with cystic fibrosis that underwent lung transplantation, and compared them to the reference, nonclinical FGSC A4 strain. Both CIs presented increased growth in comparison to that of the reference strain in the presence of physiologically relevant carbon sources. Metabolomic analyses showed that the three strains are metabolically very different from each other in these carbon sources. Furthermore, the CIs were highly susceptible to cell wall-perturbing agents but not to other physiologically relevant stresses. Genome analyses identified several frameshift variants in genes encoding cell wall integrity (CWI) signaling components. Significant differences in CWI signaling were confirmed by Western blotting among the three strains. In vivo virulence studies using several different models revealed that strain MO80069 had significantly higher virulence in hosts with impaired neutrophil function than the other strains. In summary, this study presents detailed biological characterization of two A. nidulanssensu stricto clinical isolates. Just as in Aspergillus fumigatus, strain heterogeneity exists in A. nidulans clinical strains that can define virulence traits. Further studies are required to fully characterize A. nidulans strain-specific virulence traits and pathogenicity.IMPORTANCE Immunocompromised patients are susceptible to infections with opportunistic filamentous fungi from the genus Aspergillus Although A. fumigatus is the main etiological agent of Aspergillus species-related infections, other species, such as A. nidulans, are prevalent in a condition-specific manner. A. nidulans is a predominant infective agent in patients suffering from chronic granulomatous disease (CGD). A. nidulans isolates have mainly been studied in the context of CGD although infection with A. nidulans also occurs in non-CGD patients. This study carried out a detailed biological characterization of two non-CGD A. nidulans clinical isolates and compared the results to those with a reference strain. Phenotypic, metabolomic, and genomic analyses highlight fundamental differences in carbon source utilization, stress responses, and maintenance of cell wall integrity among the strains. One clinical strain had increased virulence in models with impaired neutrophil function. Just as in A. fumigatus, strain heterogeneity exists in A. nidulans clinical strains that can define virulence traits.

Dataset Information

Molecular characterization of an adaptive response to alkylating agents in the opportunistic pathogen Aspergillus fumigatus.

Publications

Molecular characterization of an adaptive response to alkylating agents in the opportunistic pathogen Aspergillus fumigatus.

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