Illumina identification of RsrA, a conserved C2H2 transcription factor coordinating the NapA mediated oxidative stress signaling pathway in Aspergillus

Bok, Jin Woo; Wiemann, Philipp; Garvey, Graeme S.; Lim, Fang Yun; Haas, Brian J.; Wortman, Jennifer; Keller, Nancy P.

doi:10.1186/1471-2164-15-1011

Cited by 21 publications

(20 citation statements)

References 70 publications

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“…34 Briefly, 5 μ L of freshly grown (7 days old) 1 × 10 5 spores mL −1 of each strain were inoculated onto solid GMM, which had been supplemented with H 2 O 2 (5 μ M, 10 μ M, or 15 μ M), tertiary butyl hydroperoxide (tBOOH; 0.6 mM, 1.2 mM, or 1.8 mM), or menadione (20 μ M, 30 μ M, or 40 μ M). GMM without any stressor served as a control.…”

Section: Methodsmentioning

confidence: 99%

NRPS-Derived Isoquinolines and Lipopetides Mediate Antagonism between Plant Pathogenic Fungi and Bacteria

et al. 2017

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Bacterial–fungal interactions are presumed to be mediated chiefly by small-molecule signals; however, little is known about the signaling networks that regulate antagonistic relationships between pathogens. Here, we show that the ralstonins, lipopeptides produced by the plant pathogenic bacteria Ralstonia solanacearum, interfere with germination of the plant-pathogenic fungus Aspergillus flavus by down-regulating expression of a cryptic biosynthetic gene cluster (BGC), named imq. Comparative metabolomic analysis of overexpression strains of the transcription factor ImqK revealed imq-dependent production of a family of tripeptide-derived alkaloids, the imizoquins. These alkaloids are produced via a nonribosomal peptide synthetase- (NRPS-)derived tripeptide and contain an unprecedented tricyclic imidazo[2,1-a]isoquinoline ring system. We show that the imizoquins serve a protective role against oxidative stress that is essential for normal A. flavus germination. Supplementation of purified imizoquins restored wildtype germination to a ΔimqK A. flavus strain and protected the fungus from ROS damage. Whereas the bacterial ralstonins retarded A. flavus germination and suppressed expression of the imq cluster, the fungal imizoquins in turn suppressed growth of R. solanacearum. We suggest such reciprocal small-molecule-mediated antagonism is a common feature in microbial encounters affecting pathogenicity and survival of the involved species.

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Section: Methodsmentioning

confidence: 99%

NRPS-Derived Isoquinolines and Lipopetides Mediate Antagonism between Plant Pathogenic Fungi and Bacteria

et al. 2017

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“…Two independent isolates of MRB234 were sequenced on the Illumina GAIIx platform at the University of Wisconsin Biotechnology Center. MRB234 variants were detected as previously described (31). The remaining 16 MRB mutants and a nearly isogenic wild-type control were sequenced on an Ion Torrent Personal Genome Machine (PGM).…”

Section: Methodsmentioning

confidence: 99%

“…Each MRB strain was backcrossed to the same strain (RJW2) two to seven times and genomic DNA was extracted from each isolate (31). Two independent isolates of MRB234 were sequenced on the Illumina GAIIx platform at the University of Wisconsin Biotechnology Center.…”

Section: Methodsmentioning

confidence: 99%

“…These uncharacterized MRB mutants were cryogenically stored, awaiting a faster and more economical strategy for identification. Whole-genome sequencing has been used to identify mutant alleles in multiple model organisms (27)(28)(29), including several fungi (30)(31)(32)(33)(34)(35). Here we describe our success in utilizing next-generation sequencing of mutants using Illumina and Ion Torrent platforms to quickly and effectively identify the genetic basis of 12 of the remaining MRB mutants.…”

Section: Importancementioning

confidence: 99%

“…With the relatively small size of most fungal genomes and the continually declining price of whole-genome sequencing, there is now an opportunity for the revitalization of forward genetic screening for identifying and characterization of novel genetic pathways. Next-generation sequencing represents a solution to old genetic problems and can be used to move the field toward a more complete picture of gene regulation, biochemistry, and cellular biology (31,55,56). The use of Ion Torrent and Illumina sequencing technology allowed us to generate whole-genome sequences and successfully identify the causative mutations in 12 of the 17 isolates that we sequenced.…”

Section: Fig 2 Gene Deletions Of Novel Regulators Inmentioning

confidence: 99%

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Revitalization of a Forward Genetic Screen Identifies Three New Regulators of Fungal Secondary Metabolism in the Genus Aspergillus

Pfannenstiel

Zhao

Wortman

et al. 2017

mBio

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The study of aflatoxin in Aspergillus spp. has garnered the attention of many researchers due to aflatoxin's carcinogenic properties and frequency as a food and feed contaminant. Significant progress has been made by utilizing the model organism Aspergillus nidulans to characterize the regulation of sterigmatocystin (ST), the penultimate precursor of aflatoxin. A previous forward genetic screen identified 23 A. nidulans mutants involved in regulating ST production. Six mutants were characterized from this screen using classical mapping (five mutations in mcsA) and complementation with a cosmid library (one mutation in laeA). The remaining mutants were backcrossed and sequenced using Illumina and Ion Torrent sequencing platforms. All but one mutant contained one or more sequence variants in predicted open reading frames. Deletion of these genes resulted in identification of mutant alleles responsible for the loss of ST production in 12 of the 17 remaining mutants. Eight of these mutations were in genes already known to affect ST synthesis (laeA, mcsA, fluG, and stcA), while the remaining four mutations (in laeB, sntB, and hamI) were in previously uncharacterized genes not known to be involved in ST production. Deletion of laeB, sntB, and hamI in A. flavus results in loss of aflatoxin production, confirming that these regulators are conserved in the aflatoxigenic aspergilli. This report highlights the multifaceted regulatory mechanisms governing secondary metabolism in Aspergillus. Additionally, these data contribute to the increasing number of studies showing that forward genetic screens of fungi coupled with wholegenome resequencing is a robust and cost-effective technique.IMPORTANCE In a postgenomic world, reverse genetic approaches have displaced their forward genetic counterparts. The techniques used in forward genetics to identify loci of interest were typically very cumbersome and time-consuming, relying on Mendelian traits in model organisms. The current work was pursued not only to identify alleles involved in regulation of secondary metabolism but also to demonstrate a return to forward genetics to track phenotypes and to discover genetic pathways that could not be predicted through a reverse genetics approach. While identification of mutant alleles from whole-genome sequencing has been done before, here we illustrate the possibility of coupling this strategy with a genetic screen

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The bZIP-type transcription factors NapA and RsmA modulate the volumetric ratio and the relative superoxide ratio of mitochondria in Aspergillus nidulans

Bákány,

Antal,

Szentesi

et al. 2023

BIOLOGIA FUTURA

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Basic leucine zipper (bZIP) transcription factors are crucial components of differentiation, cellular homeostasis and the environmental stress defense of eukaryotes. In this work, we further studied the consequence of gene deletion and overexpression of two bZIP transcription factors, NapA and RsmA, on superoxide production, mitochondrial morphology and hyphal diameter of Aspergillus nidulans. We have found that reactive oxygen species production was influenced by both gene deletion and overexpression of napA under tert-butylhydroperoxide (tBOOH) elicited oxidative stress. Furthermore, gene expression of napA negatively correlated with mitochondrial volumetric ratio as well as sterigmatocystin production of A. nidulans. High rsmA expression was accompanied with elevated relative superoxide ratio in the second hyphal compartment. A negative correlation between the expression of rsmA and catalase enzyme activity or mitochondrial volumetric ratio was also confirmed by statistical analysis. Hyphal diameter was independent on either rsmA and napA expression as well as 0.2 mM tBOOH treatment.

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Illumina identification of RsrA, a conserved C2H2 transcription factor coordinating the NapA mediated oxidative stress signaling pathway in Aspergillus

Cited by 21 publications

References 70 publications

NRPS-Derived Isoquinolines and Lipopetides Mediate Antagonism between Plant Pathogenic Fungi and Bacteria

NRPS-Derived Isoquinolines and Lipopetides Mediate Antagonism between Plant Pathogenic Fungi and Bacteria

Revitalization of a Forward Genetic Screen Identifies Three New Regulators of Fungal Secondary Metabolism in the Genus Aspergillus

The bZIP-type transcription factors NapA and RsmA modulate the volumetric ratio and the relative superoxide ratio of mitochondria in Aspergillus nidulans

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