Cytochrome P450s (CYPs), heme-containing monooxygenases, play important roles in a wide variety of metabolic processes important for development as well as biotic/trophic interactions in most living ...organisms. Functions of some CYP enzymes are similar across organisms, but some are organism-specific; they are involved in the biosynthesis of structural components, signaling networks, secondary metabolisms, and xenobiotic/drug detoxification. Fungi possess more diverse CYP families than plants, animals, or bacteria. Various fungal CYPs are involved in not only ergosterol synthesis and virulence but also in the production of a wide array of secondary metabolites, which exert toxic effects on humans and other animals. Although few studies have investigated the functions of fungal CYPs, a recent systematic functional analysis of CYP genes in the plant pathogen
identified several novel CYPs specifically involved in virulence, asexual and sexual development, and degradation of xenobiotics. This review provides fundamental information on fungal CYPs and a new platform for further metabolomic and biochemical studies of CYPs in toxigenic fungi.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Abstract
Alternaria
is an important toxigenic fungal genus endangering the quality of cereal grains. No data are available in the literature regarding the composition of
Alternaria
species in wheat ...from the main wheat‐cultivating regions of China. In this study, a total of 282 strains were isolated from wheat kernels collected in various provinces. Phylogenetic characterization was performed using two informative loci,
ASA10
and
ASA19
, demonstrating that all strains belonged to
A
.
alternata
. A low molecular diversity was observed, which did not appear to be influenced by geographic origin. The secondary metabolite profile of the strains was evaluated in rice culture. Altertoxin I was synthesized at a high concentration by the majority of the strains, while the accumulation of altenuene was lower in both frequency and level than that of alternariol and its derivative.
A
.
alternata
did not cause readily observable symptoms on wheat spikes after spraying of the conidia at anthesis, but a higher quantity of altertoxin I and
Alternaria
DNA were detected in inoculated grains. Artificial inoculations of apple fruit showed that all examined strains were capable of causing typical rot symptoms and producing all four toxins to a certain degree. The widespread occurrence of
A
.
alternata
on wheat grains and its ability to synthesize various toxic metabolites may pose a serious hazard for both growers and consumers.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Summary
The role of AtMYB44, an R2R3 MYB transcription factor, in signaling mediated by jasmonic acid (JA) and salicylic acid (SA) is examined. AtMYB44 is induced by JA through CORONATINE ...INSENSITIVE 1 (COI1). AtMYB44 over‐expression down‐regulated defense responses against the necrotrophic pathogen Alternaria brassicicola, but up‐regulated WRKY70 and PR genes, leading to enhanced resistance to the biotrophic pathogen Pseudomonas syringae pv. tomato DC3000. The knockout mutant atmyb44 shows opposite effects. Induction of WRKY70 by SA is reduced in atmyb44 and npr1‐1 mutants, and is totally abolished in atmyb44 npr1‐1 double mutants, showing that WRKY70 is regulated independently through both NPR1 and AtMYB44. AtMYB44 over‐expression does not change SA content, but AtMYB44 over‐expression phenotypes, such as retarded growth, up‐regulated PR1 and down‐regulated PDF1.2 are reversed by SA depletion. The wrky70 mutation suppressed AtMYB44 over‐expression phenotypes, including up‐regulation of PR1 expression and down‐regulation of PDF1.2 expression. β‐estradiol‐induced expression of AtMYB44 led to WRKY70 activation and thus PR1 activation. AtMYB44 binds to the WRKY70 promoter region, indicating that AtMYB44 acts as a transcriptional activator of WRKY70 by directly binding to a conserved sequence element in the WRKY70 promoter. These results demonstrate that AtMYB44 modulates antagonistic interaction by activating SA‐mediated defenses and repressing JA‐mediated defenses through direct control of WRKY70.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Deoxynivalenol (DON) is commonly found in wheat and wheat-derived foods, posing a threat to human health. Biodegradation is an efficient and eco-friendly measure for mycotoxin detoxification. ...Understanding the mechanism of DON biodegradation is hence of great importance. Herein, we report the application of metabolomics methods for the analysis of DON degradation by a bacterial consortium isolated from wheat leaves collected in Jiangsu Province. Metabolomics analysis combined with a nuclear magnetic resonance analysis revealed the main degradation product, 3-keto-DON, and a minor degradation product, 3-epi-DON. Further study illustrated that DON underwent a two-step epimerization through the intermediate 3-keto-DON. Sequencing analysis of the 16S rRNA metagenome of the microorganismal community suggested that the abundance of three bacterial genera,
Achromobacter
,
Sphingopyxis
, and
Sphingomonas
, substantially increased during the coculture of bacterial consortium and DON. Further investigation revealed that
Devosia
sp. might be responsible for the epimerization of 3-keto-DON. These findings shed light on the catabolic pathways of DON during biodegradation and illustrate the potential of using metabolomics approaches in biodegradation studies.
Key Points
• A bacterial consortium was isolated with good deoxynivalenol-degrading potential.
• Metabolomics approaches were successfully used to interpret the degradation pathway.
• A trace-amount degradation product was determined by metabolomics and NMR analysis.
Graphical Abstract
.
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CEKLJ, DOBA, EMUNI, FZAB, GEOZS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NUK, OBVAL, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Summary
Fungal sexual reproduction requires complex cellular differentiation processes of hyphal cells. The plant pathogenic fungus Fusarium graminearum produces fruiting bodies called perithecia via ...sexual reproduction, and perithecia forcibly discharge ascospores into the air for disease initiation and propagation. Lipid metabolism and accumulation are closely related to perithecium formation, yet the molecular mechanisms that regulate these processes are largely unknown. Here, we report that a novel fungal specific bZIP transcription factor, F. graminearum perithecium overproducing 1 (Fpo1), plays a role as a global transcriptional repressor during perithecium production and maturation in F. graminearum. Deletion of FPO1 resulted in reduced vegetative growth, asexual sporulation and virulence and overproduced perithecium, which reached maturity earlier, compared with the wild type. Intriguingly, the hyphae of the fpo1 mutant accumulated excess lipids during perithecium production. Using a combination of molecular biological, transcriptomic and biochemical approaches, we demonstrate that repression of FPO1 after sexual induction leads to reprogramming of carbon metabolism, particularly fatty acid production, which affects sexual reproduction of this fungus. This is the first report of a perithecium‐overproducing F. graminearum mutant, and the findings provide comprehensive insight into the role of modulation of carbon metabolism in the sexual reproduction of fungi.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Various ascomycete fungi possess sex-specific molecular mechanisms, such as repeat-induced point mutations, meiotic silencing by unpaired DNA, and unusual adenosine-to-inosine RNA editing, for genome ...defense or gene regulation. Using a combined analysis of functional genetics and deep sequencing of small noncoding RNA (sRNA), mRNA, and the degradome, we found that the sex-specifically induced exonic small interference RNA (ex-siRNA)-mediated RNA interference (RNAi) mechanism has an important role in fine-tuning the transcriptome during ascospore formation in the head blight fungus Fusarium graminearum. Approximately one-third of the total sRNAs were produced from the gene region, and sRNAs with an antisense direction or 5'-U were involved in post-transcriptional gene regulation by reducing the stability of the corresponding gene transcripts. Although both Dicers and Argonautes partially share their functions, the sex-specific RNAi pathway is primarily mediated by FgDicer1 and FgAgo2, while the constitutively expressed RNAi components FgDicer2 and FgAgo1 are responsible for hairpin-induced RNAi. Based on our results, we concluded that F. graminearum primarily utilizes ex-siRNA-mediated RNAi for ascosporogenesis but not for genome defenses and other developmental stages. Each fungal species appears to have evolved RNAi-based gene regulation for specific developmental stages or stress responses. This study provides new insights into the regulatory role of sRNAs in fungi and other lower eukaryotes.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Summary
Reactive oxygen species (ROS) are associated with various developmental processes and host–pathogen interactions in pathogenic fungi. Peroxidases are a group of ROS‐detoxifying enzymes that ...are involved in the oxidative stress response and in a variety of physiological processes. In this study, we performed a genome‐wide functional characterization of putative peroxidase genes in Fusarium graminearum, a head blight pathogen of cereal crops. We identified 31 putative peroxidase genes and generated deletion mutants for these genes. Twenty‐six of the deletion mutants showed developmental phenotypes indistinguishable from that of the wild‐type, and five deletion mutants exhibited phenotypic changes in at least one phenotypic category. Four deletion mutants, fca6, fca7, fpx1 and fpx15, showed increased sensitivity to extracellular H2O2. Deletion mutants of FCA7 also exhibited reduced virulence and increased trichothecene production compared with those of the wild‐type strain, suggesting that Fca7 may play an important role in the host–pathogen interaction in F. graminearum. To identify the transcription factors (TFs) regulating FCA6, FCA7, FPX1 and FPX15 in response to oxidative stress, we screened an F. graminearum TF mutant library for growth in the presence of H2O2 and found that multiple TFs co‐regulated the expression of FCA7 under oxidative stress conditions. These results demonstrate that a complex network of transcriptional regulators of antioxidant genes is involved in oxidative stress responses in this fungus. Moreover, our study provides insights into the roles of peroxidases in developmental processes and host–pathogen interactions in plant‐pathogenic fungi.
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BFBNIB, DOBA, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, UILJ, UKNU, UL, UM, UPUK
Many fungi-infecting viruses, which are termed mycoviruses, have been identified, and most do not cause any visible symptoms. Some mycoviruses, however, can attenuate the virulence of the infected ...fungi, a phenomenon referred to as hypovirulence. To study fungus responses to virus infection, we established a model system composed of Fusarium graminearum and four mycoviruses including FgV1 (Fusarium graminearum virus 1), FgV2, FgV3, and FgV4. FgV1 and FgV2 infections caused several phenotypic alterations in F. graminearum including abnormal colony morphology, defects in perithecium development, and reductions in growth rate, conidiation, and virulence. In contrast, FgV3 and FgV4 infections did not cause any phenotypic change. An RNA-Seq-based analysis of the host transcriptome identified four unique Fusarium transcriptomes, one for each of the four mycoviruses. Unexpectedly, the fungal host transcriptome was more affected by FgV1 and FgV4 infections than by FgV2 and FgV3 infections. Gene ontology (GO) enrichment analysis revealed that FgV1 and FgV3 infections resulted in down-regulation of host genes required for cellular transport systems. FgV4 infection reduced the expression of genes involved in RNA processing and ribosome assembly. We also found 12 genes that were differentially expressed in response to all four mycovirus infections. Unfortunately, functions of most of these genes are still unknown. Taken together, our analysis provides further detailed insights into the interactions between mycoviruses and F. graminearum.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Aims
Cereals contaminated with type B trichothecene nivalenol (NIV) and its acetylated derivative 4‐acetyl‐nivalenol (4‐AcNIV) are a global mycotoxicological problem threatening the health of humans ...and livestock. Toxicological studies, quantitative determinations and screening for biodegrading micro‐organisms require massive amounts of pure toxins. However, the low yield from fungal cultures and high prices of NIV and 4‐AcNIV limit research progress in these areas. This work aimed to select Fusarium asiaticum mutant strains with enhanced production of NIV and 4‐AcNIV.
Methods and Results
A total of 62 NIV‐producing F. asiaticum strains were isolated and compared regarding their ability to produce NIV. Strain RR108 had the highest yield of NIV among 62 field isolates surveyed and was then genetically modified for higher production. Targeted deletion of the FaFlbA gene, encoding a regulator of G protein signalling protein, resulted in a significant increase in NIV and 4‐AcNIV production in the FaFlbA deletion mutant ΔFaFlbA. The expression of three TRI genes involved in the trichothecene biosynthetic pathway was upregulated in ΔFaFlbA. ΔFaFlbA produced the highest amount of NIV and 4‐AcNIV when cultured in brown long‐grain rice for 21 days, and the yields were 2.07 and 2.84 g kg−1, respectively. The mutant showed reduced fitness, including reduced conidiation, loss of perithecial development and decreased virulence on wheat heads, which makes it biologically safe for large‐scale preparation and purification of NIV and 4‐AcNIV.
Conclusions
The F. asiaticum mutant strain ΔFaFlbA presented improved production of NIV and 4‐AcNIV with reduced fitness and virulence in plants.
Significance and Impact of the Study
Targeted deletion of the FaFlbA gene resulted in increased NIV and 4‐AcNIV production. Our results provide a practical approach using genetic modification for large‐scale mycotoxin production.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Deoxynivalenol (DON), a common mycotoxin of type B trichothecene, is produced mainly by several
species. DON causes great losses in farming and poses severe safety risks to human and animal health. ...Thus, DON contamination in cereals and DON toxicity are of worldwide concern. In this study, we screened the bacterial consortium C20, which efficiently degraded almost 70 μg ml
DON within 5 days. The bacterial consortium also had the ability to degrade 15-acetyl-DON, 3-acetyl-DON, and T-2 toxin. The bacterial consortium C20 was able to degrade DON under a wide range of pH and temperature conditions. The optimal temperature and pH for DON degradation were 30°C and pH 8.0, respectively. The bacterial consortium C20 comprised of different bacterial genera, and several strains were found to significantly increase when cultured in Mineral Medium with 100 μg ml
DON based on the analysis of the sequences of the hypervariable V3-V4 region of the 16S rRNA gene. 3-keto-DON was confirmed as a degradation product of DON by liquid chromatography/time-of-flight/mass spectrometry (LC-TOF-MS) and nuclear magnetic resonance (NMR) analyses. The results indicated that the bacterial consortium C20 is a potential candidate for the biodegradation of DON in a safe and environmentally friendly manner.
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