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Biological functions of the bZIP transcription factor FgMetR in Fusarium graminearum
MA Hao, ZHANG Limin, ZHAO Yanxiang, HUANG Jinguang
Mycosystema ›› 2025, Vol. 44 ›› Issue (4) : 240279.
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Biological functions of the bZIP transcription factor FgMetR in Fusarium graminearum
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Fusarium head blight (FHB) caused by Fusarium graminearum is one of the most important fungal diseases of wheat, posing a serious threat to wheat production and food safety. bZIP transcription factors are core positive regulators of sulfur assimilation and involved in the regulation of fungal asexual reproduction and pathogenicity. In order to further reveal the biological function of the bZIP transcription factors in F. graminearum, in this study, the authors identified a bZIP transcription factor FgMetR in F. graminearum and obtained the deletion mutant ΔFgMetR. The effect of FgMetR deletion on various phenotype was evaluated. The results showed that the colony of ΔFgMetR appeared transparent on PDA plates and the mycelia were sparsely dispersed with few aerial hyphae. No red pigment was observed either on PDA plates or in PDB broth. The radial growth rate of colonies, mycelial height, and conidial production of ΔFgMetR were significantly reduced as compared with those of the wild-type strain. The growth defect phenotype of ΔFgMetR on minimal medium without any sulfur element can be partially restored by supplementing organic but not inorganic sulfur sources. ΔFgMetR mutant failed to form perithecia and ascospores on carrot culture medium. Deletion of FgMetR significantly reduced the pathogenicity of F. graminearum invading both wheat coleoptiles and heads. In addition, ΔFgMetR mutant was more sensitive to tebuconazole and prochloraz. The facts prove that bZIP transcription factor FgMetR is essential for the vegetative growth, pigmentation, asexual and sexual reproduction, pathogenicity and involvement in the regulation of sulfur assimilation and sensitivity to azole fungicides of F. graminearum.
Fusarium head blight / transcriptional regulation / sulfur assimilation / pathogenicity / azole fungicide {{custom_keyword}} /
Table 1 Content of main active components in CME表1 CME中主要活性成分含量 |
| 成分 Components | 含量 Content (mg/g) |
|---|---|
| 多糖 Polysaccharide | 509.47±14.73 |
| 尿苷Uridine | 1.21±0.10 |
| 鸟苷Guanosine | 0.66±0.03 |
| 腺苷Adenosine | 1.47±0.11 |
| 虫草素 Cordycepin | 2.66±0.15 |
| N6-(2-羟乙基)腺苷 N6-(2-hydroxyethtl)-adenosine | 2.15±0.09 |
| 总多酚 Total polyphenols | 9.21±0.37 |
| 类胡萝卜素 Carotenoid | 0.95±0.04 |
Fig. 1 Determination of monosaccharide composition by HPLC. A: Mixed reference substance; B: Tested samples. 1: Man; 2: Rha; 3: Glc-UA; 4: Gal-UA; 5: Lactose (internal standard); 6: Glc; 7: Gal: 8: Xyl; 9: Ara; 10: Fuc.图1 HPLC检测单糖组成 A：混合对照品；B：供试品. 1：甘露糖；2：鼠李糖；3：葡萄糖醛酸；4：半乳糖醛酸；5：乳糖内标；6：葡萄糖；7：半乳糖；8：木糖；9：阿拉伯糖；10：岩藻糖 |
Table 2 Monosaccharide composition of Cordyceps militaris polysaccharide表2 蛹虫草多糖的单糖组成 |
| 样品 Sample | 甘露糖 Man | 半乳糖醛酸 Gal-UA | 葡萄糖 Glc | 半乳糖 Gal | 阿拉伯糖 Ara |
|---|---|---|---|---|---|
| 含量 Content (mol%) | 25.73 | 9.26 | 29.01 | 29.39 | 5.62 |
Fig. 2 Determination of nucleosides by HPLC. A: Mixed reference substance; B: Test sample. 1: Cytidine; 2: Uridine; 3: Guanosine; 4: Thymidine; 5: Adenosine; 6: Cordycepin; 7: N6-(2-hydroxyethtl)- adenosine.图2 HPLC检测核苷类成分 A：混合对照品；B：供试品. 1：胞苷；2：尿苷；3：鸟苷；4：胸苷；5：腺苷；6：虫草素；7：N6-(2-羟乙基)腺苷 |
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The mycotoxin deoxynivalenol (DON), produced by several Fusarium spp., acts as a virulence factor and is essential for symptom development after initial wheat infection. Accumulating evidence shows that the production of this secondary metabolite can be triggered by diverse environmental and cellular signals, implying that it might have additional roles during the life cycle of the fungus. Here, we review data that position DON in the saprophytic fitness of Fusarium, in defense and in the primary C and N metabolism of the plant and the fungus. We combine the available information in speculative models on the role of DON throughout the interaction with the host, providing working hypotheses that await experimental validation. We also highlight the possible impact of control measures in the field on DON production and summarize the influence of abiotic factors during processing and storage of food and feed matrices. Altogether, we can conclude that DON is a very important compound for Fusarium to cope with a changing environment and to assure its growth, survival, and production of toxic metabolites in diverse situations.
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Expression of the sulfur assimilation pathway in Aspergillus nidulans is under control of sulfur metabolite repression, which is composed of scon genes encoding subunits of ubiquitin ligase and the metR gene coding for a transcriptional activator. In this paper we report three dominant suppressors of methionine requirement isolated from a metB3 diploid strain. All three mutations lead to the substitution of phenylalanine 48 by serine or leucine in the conserved N-terminal region of the MetR protein. Strains carrying the dominant suppressor mutations exhibit increased activities of homocysteine synthase and sulfur assimilation enzymes as well as elevated levels of the corresponding transcripts. These changes are observed even under conditions of methionine repression, which suggests that the mutated MetR protein may be resistant to inactivation or degradation mediated by sulfur metabolite repression. We also found that a mutant impaired in sulfite reductase activity, known until now as sG8, has a frameshift which changes 41 C-terminal amino acids. Therefore, it is now designated metR18. This mutant has elevated levels of MetR-regulated transcripts and of activities of sulfur assimilation enzymes (except sulfite reductase), which can be repressed to the wild type level by exogenous methionine. Thus, metR18 and the three dominant suppressors represent new types of mutations affecting different parts of the A. nidulans MetR protein.Copyright © 2010 Elsevier Inc. All rights reserved.
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Fusarium head blight (FHB) of small grain cereals caused by and other species is an economically important plant disease worldwide. infections not only result in severe yield losses but also contaminate grain with various mycotoxins, especially deoxynivalenol (DON). With the complete genome sequencing of, tremendous progress has been made during the past two decades toward understanding the basis for DON biosynthesis and its regulation. Here, we summarize the current understanding of DON biosynthesis and the effect of regulators, signal transduction pathways, and epigenetic modifications on DON production and the expression of biosynthetic genes. In addition, strategies for controlling FHB and DON contamination are reviewed. Further studies on these biosynthetic and regulatory systems will provide useful knowledge for developing novel management strategies to prevent FHB incidence and mycotoxin accumulation in cereals.
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Saccharomyces cerevisiae contains eight members of a novel and fungus-specific family of bZIP proteins that is defined by four atypical residues on the DNA-binding surface. Two of these proteins, Yap1 and Yap2, are transcriptional activators involved in pleiotropic drug resistance. Although initially described as AP-1 factors, at least four Yap proteins bind most efficiently to TTACTAA, a sequence that differs at position +/-2 from the optimal AP-1 site (TGACTCA); further, a Yap-like derivative of the AP-1 factor Gcn4 (A239Q S242F) binds efficiently to the Yap recognition sequence. Molecular modeling suggests that the Yap-specific residues make novel contacts and cause physical constraints at the +/-2 position that may account for the distinct DNA-binding specificities of Yap and AP-1 proteins. To various extents, Yap1, Yap2, Yap3, and Yap5 activate transcription from a promoter containing a Yap recognition site. Yap-dependent transcription is abolished in strains containing high levels of protein kinase A; in contrast, Gcn4 transcriptional activity is stimulated by protein kinase A. Interestingly, Yap1 transcriptional activity is stimulated by hydrogen peroxide, whereas Yap2 activity is stimulated by aminotriazole and cadmium. In addition, unlike other yap mutations tested, yap4 (cin5) mutations affect chromosome stability, and they suppress the cold-sensitive phenotype of yap1 mutant strains. Thus, members of the Yap family carry out overlapping but distinct biological functions.
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Fusarium head blight (FHB) is a devastating disease of cereal crops worldwide mainly caused by Fusarium graminearum. Due to the unavailability of FHB-resistant wheat cultivars, chemical fungicide application is currently the most effective approach for controlling FHB now. In the last few years, a novel cyanoacrylate fungicide, phenamacril, has been widely used in China for FHB disease management. In previous studies, we identified that myosin I (FgMyo1) is the target of phenamacril and is essential for mycotoxin deoxynivalenol (DON) biosynthesis and fungal growth. However, the regulation of FgMYO1 gene expression is still largely unknown.In this study, we identified a b-ZIP transcription factor, FgTfmI, which regulates the mRNA expression of FgMYO1 upon phenamacril treatment. The FgTfmI directly binds to the promoter region of FgMYO1, and is required for the upregulation of FgMYO1 in response to phenamacril treatment. The deletion mutant of FgTFMI (ΔFgTfmI) displayed a slight growth defect, while it showed hypersensitivity to phenamacril, but not to other tested fungicides. FgTfmI also contributed to DON biosynthesis and the infection process in planta.The transcription factor FgTfmI plays an important role in regulating transcription of the genes involved in phenamacril tolerance, DON biosynthesis and virulence in F. graminearum. © 2019 Society of Chemical Industry.© 2019 Society of Chemical Industry.
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A met4 mutant of Saccharomyces cerevisiae was unable to transcribe a number of genes encoding enzymes of the methionine biosynthetic pathway. The sequence of the cloned MET4 gene allowed the previously sequenced flanking LEU4 and POL1 genes to be linked to MET4 into a 10,327 bp contiguous region of chromosome XIV. From the sequence and mapping of the transcriptional start points, MET4 is predicted to encode a protein of 634 amino acids (as opposed to 666 amino acids published by others) with a leucine zipper domain at the C-terminus, preceded by both acidic and basic regions. Thus, MET4 belongs to the family of basic leucine zipper trans-activator proteins. Disruption of MET4 resulted in methionine auxotrophy with no other phenotype. Transcriptional studies showed that MET4 was regulated by the general amino acid control and hence by another bZIP protein encoded by GCN4. GCN4 binding sequences are present between the divergently transcribed MET4 and LEU4 genes. Over-expression of MET4 resulted in leaky expression from the otherwise tightly regulated MET3 promoter under its control. The presence of consensus sequences for other potential regulatory elements in the MET4 promoter suggests a complex regulation of this gene.
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The identification, isolation and characterization of a new Aspergillus nidulans positive-acting gene metR, which encodes a transcriptional activator of sulphur metabolism, is reported. metR mutants are tight auxotrophs requiring methionine or homocysteine for growth. Mutations in the metR gene are epistatic to mutations in the negative-acting sulphur regulatory scon genes. The metR coding sequence is interrupted by a single intron of 492 bp which is unusually long for fungi. Aspergillus nidulans METR is a member of bZIP family of DNA-binding proteins. The bZIP domains of METR and the Neurospora crassa CYS3 transcriptional activator of sulphur genes are highly similar. Although Neurospora cys-3 gene does not substitute for the metR function, a chimeric metR gene with a cys-3 bZIP domain is able to transform the DeltametR mutant to methionine prototrophy. This indicates that METR recognizes the same regulatory sequence as CYS3. The metR gene is not essential, as deletion mutants are viable and have similar phenotype as point mutants. In contrast to the Neurospora cys-3, transcription of the metR gene was found to be regulated neither by METR protein nor by sulphur source. Transcription of metR gene is derepressed in the sconB2 mutant. Transcription of genes encoding sulphate permease, homocysteine synthase, cysteine synthase, ATP-sulphurylase, and sulphur controller--sconB is strongly regulated by the metR gene product and depends on the character of the metR mutation and sulphur supplementation.
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The CYS3 transcription factor is a basic region-leucine zipper (bZIP) DNA-binding protein that is essential for the expression of a coordinately regulated group of genes involved in the acquisition and utilization of sulfur in Neurospora crassa. An approach of using binding-site selection from random-sequence oligonucleotides was used to define CYS3-binding specificity. The derived consensus-binding site of ATGGCGCCAT defines a symmetrical sequence (half-site A T G/t G/a C/t) that resembles that of other bZIP proteins such as CREB and C/EBP. By comparison, CYS3 shows a greater range of binding to a central core of varied Pur-Pyr-Pur-Pyr sequences than CREB as determined by gel shift assays. The derived CYS3 consensus binding sequence was further validated by demonstrating in vivo sulfur regulation using a heterologous promoter construct. The CYS3-binding site data will be useful for the genome-wide study of sulfur-regulated genes in N. crassa, which has served as a model fungal sulfur control system.
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'The gastrointestinal tract represents the first barrier against food contaminants as well as the first target for these toxicants. Deoxynivalenol (DON) is a mycotoxin that commonly contaminates cereals and causes various toxicological effects. Through consumption of contaminated cereals and cereal products, human and pigs are exposed to this mycotoxin. Using in vitro, ex vivo and in vivo approaches, we investigated the effects of DON on the intestinal epithelium. We demonstrated that, in intestinal epithelial cell lines from porcine (IPEC-1) or human (Caco-2) origin, DON decreases trans-epithelial electrical resistance (TEER) and increases in a time and dose-dependent manner the paracellular permeability to 4 kDa dextran and to pathogenic Escherichia coli across intestinal cell monolayers. In pig explants treated with DON, we also observed an increased permeability of intestinal tissue. These alterations of barrier function were associated with a specific reduction in the expression of claudins, which was also seen in vivo in the jejunum of piglets exposed to DON-contaminated feed. In conclusion, DON alters claudin expression and decreases the barrier function of the intestinal epithelium. Considering that high levels of DON may be present in food or feed, consumption of DON-contaminated food/feed may induce intestinal damage and has consequences for human and animal health.
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Mould growth and mycotoxin production are related to plant stress caused by environmental factors such as: extreme weather; insect damage; inadequate storage conditions and incorrect fertilization; these predispose plants to mycotoxin contamination in the field. Fusarium species infect wheat during the flowering period. In addition to losses of yield, these fungi can also synthesize toxic components (mycotoxins) in suitable environmental conditions, thus threatening animal and human health. Given the severe consequences and the fact that mycotoxins affect production throughout the world, the ability to predict Fusarium head blight (FHB) and deoxynivalenol (DON) and other mycotoxin contamination is important to reduce the year-to-year risk for producers. Owing to these dangerous consequences in Argentina, Belgium, Canada, Italy, the United States and in Europe, computer models, based on weather variables (temperature, rainfall and moisture level), have been developed to predict the occurrence of FHB and DON contamination in wheat.
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The production of trichothecene mycotoxins by some plant pathogenic species of Fusarium is thought to contribute to their virulence. Gibberella zeae (F. graminearum) is an important cereal pathogen that produces the trichothecene deoxynivalenol. To determine if trichothecene production contributes to the virulence of G. zeae, we generated trichothecene-deficient mutants of the fungus by gene disruption. The disrupted gene, Tri5, encodes the enzyme trichodiene synthase, which catalyzes the first step in trichothecene biosynthesis. To disrupt Tri5, G. zeae was transformed with a plasmid carrying a doubly truncated copy of the Tri5 coding region interrupted by a hygromycin B resistance gene. Tri5- transformants were selected by screening for the inability to produce trichothecenes and by Southern blot analysis. Tri5- strains exhibited reduced virulence on seedlings of Wheaton wheat and common winter rye, but wild-type virulence on seedlings of Golden Bantam maize. On Caldwell and Marshall wheat and Porter oat seedlings, Tri5- strains were inconsistent in causing less disease than their wild-type progenitor strain. Head blight developed more slowly on Wheaton when inoculated with Tri5- mutants than when inoculated with wild-type strains. These results suggest that trichothecene production contributes to the virulence of G. zeae on some hosts.
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The C/EBP family of proteins represents an important group of bZIP transcription factors that are key to the regulation of essential functions such as cell cycle, hematopoiesis, skeletal development, and host immune responses. They are also intimately associated with tumorigenesis and viral disease. These proteins are regulated at multiple levels, including gene induction, alternative translational initiation, post-translational modification, and protein-protein interaction. This review attempts to integrate recent reports with more than 20 years of previous effort focused on this fascinating collection of regulators.Copyright © 2010 Elsevier Ltd. All rights reserved.
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刘礼广, 丁兆建, 李梦涵, 吴春花, 谢欣捷, 彭军, 张欣, 2023. 尖孢镰孢菌古巴专化型胱硫醚γ-合成酶的功能分析. 菌物学报, 42(7): 1575-1587
甲硫氨酸在真菌、细菌和植物的生物学过程中起着重要作用。禾谷镰刀菌Fusarium graminearum的FgMETB基因编码一个胱硫醚γ-合成酶，是甲硫氨酸合成所必需的。本研究利用同源重组的方法，在尖孢镰刀菌古巴专化型4号生理小种Fusarium oxysporum f. sp. cubense race 4 (Foc4)获得了FgMETB同源基因FoMETB的敲除突变体菌株；与野生型菌株相比，突变体菌株 ΔFoMETB 在以SO<sub>4</sub><sup>2-</sup>为唯一硫源的基本培养基(minimal medium)上不能生长。1 mmol/L甲硫氨酸的添加恢复了突变体菌株ΔFoMETB的生长，但半胱氨酸的添加不能恢复该缺失突变体的生长，说明FoMETB的敲除阻遏了Foc4半胱氨酸转化甲硫氨酸的通路。此外，ΔFoMETB的气生菌丝和菌丝干重明显减少、分枝增多、产孢量显著降低、疏水性缺失和对巴西蕉组培苗的致病性显著减弱。由此表明，FoMETB参与调控尖孢镰刀菌古巴专化型的生理特性和致病性，甲硫氨酸合成途径的关键合酶FoMETB有望成为新的抗真菌药物靶标。
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Table 1 Content of main active components in CME
Fig. 1 Determination of monosaccharide composition by HPLC. A: Mixed reference substance; B: Tested samples. 1: Man; 2: Rha; 3: Glc-UA; 4: Gal-UA; 5: Lactose (internal standard); 6: Glc; 7: Gal: 8: Xyl; 9: Ara; 10: Fuc.Table 2 Monosaccharide composition of Cordyceps militaris polysaccharideFig. 2 Determination of nucleosides by HPLC. A: Mixed reference substance; B: Test sample. 1: Cytidine; 2: Uridine; 3: Guanosine; 4: Thymidine; 5: Adenosine; 6: Cordycepin; 7: N6-(2-hydroxyethtl)- adenosine.Fig. 3 Scavenging effects of CME on radicals. A: DPPH radical scavenging activities; B: ABTS radical scavenging activities; C: ·OH radical scavenging activities.Fig. 4 Effects of different concentrations of CME on the viabilities of HaCaT cells.Fig. 5 Effects of different concentrations of H2O2 on the viabilities of HaCaT cells.Fig. 6 Effects of different concentrations of CME on the viabilities of HaCaT cells induced by H2O2. Compared with control group, ##P<0.01; Compared with model group, *P<0.05, **P<0.01. The same below.Fig. 7 Effects of different concentrations of CME on ROS levels in HaCaT cells.Fig. 8 Effects of different concentrations of CME on MDA contents (A) and the activities of SOD (B), GSH-Px (C), CAT (D) in HaCaT cells.
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