HapX-mediated H2B deub1 and SreA-mediated H2A.Z deposition coordinate in fungal iron resistance.

Plant pathogens are challenged by host-derived iron starvation or excess during infection, but the mechanism through which pathogens counteract iron stress is unclear. Here, we found that Fusarium graminearum encounters iron excess during the colonization of wheat heads. Deletion of heme activator protein X (FgHapX), siderophore transcription factor A (FgSreA) or both attenuated virulence. Further, we found that FgHapX activates iron storage under iron excess by promoting histone H2B deubiquitination (H2B deub1) at the promoter of the responsible gene. Meanwhile, FgSreA is shown to inhibit genes mediating iron acquisition during iron excess by facilitating the deposition of histone variant H2A.Z and histone 3 lysine 27 trimethylation (H3K27 me3) at the first nucleosome after the transcription start site. In addition, the monothiol glutaredoxin FgGrx4 is responsible for iron sensing and control of the transcriptional activity of FgHapX and FgSreA via modulation of their enrichment at target genes and recruitment of epigenetic regulators, respectively. Taken together, our findings elucidated the molecular mechanisms for adaptation to iron excess mediated by FgHapX and FgSreA during infection in F. graminearum and provide novel insights into regulation of iron homeostasis at the chromatin level in eukaryotes.

MAP Kinase FgHog1 and Importin ß FgNmd5 Regulate Calcium Homeostasis in Fusarium graminearum.

Maintaining cellular calcium (Ca2+) homeostasis is essential for many aspects of cellular life. The high-osmolarity glycerol (HOG) mitogen-activated protein kinase (MAPK) pathway responsible for signal integration and transduction plays crucial roles in environmental adaptation, especially in the response to osmotic stress. Hog1 is activated by transient Ca2+ increase in yeast, but the functions of the HOG pathway in Ca2+ homeostasis are largely unknown. We found that the HOG pathway was involved in the regulation of Ca2+ homeostasis in Fusarium graminearum, a devastating fungal pathogen of cereal crops. The deletion mutants of HOG pathway displayed increased sensitivity to Ca2+ and FK506, and elevated intracellular Ca2+ content. Ca2+ treatment induced the phosphorylation of FgHog1, and the phosphorylated FgHog1 was transported into the nucleus by importin ß FgNmd5. Moreover, the increased phosphorylation and nuclear accumulation of FgHog1 upon Ca2+ treatment is independent of the calcineurin pathway that is conserved and downstream of the Ca2+ signal. Taken together, this study reported the novel function of FgHog1 in the regulation of Ca2+ homeostasis in F. graminearum, which advance the understanding of the HOG pathway and the association between the HOG and calcineurin pathways in fungi.

Limonene formulation exhibited potential application in the control of mycelial growth and deoxynivalenol production in Fusarium graminearum.

Preventing grain from fungi and subsequent mycotoxins contamination has attracted notable attention. Present study demonstrated the limonene-formulated product Wetcit®, might be a biocontrol agent and potential alternative to synthetic fungicides to control Fusarium graminearum growth and deoxynivalenol (DON) production. The limonene formulation exhibited antifungal activity against F. graminearum with the EC50 at 1.40 µl/ml, electron microscopy and staining analysis showed limonene formulation could significantly decrease the quantity, length and septa of conidia, caused hyphal break and shrink, damaged the structures of cell membrane, cell wall, vacuoles and organelles in the hypha. Further study revealed the antifungal and antitoxic mechanism of limonene formulation against F. graminearum, limonene formulation significantly inhibited the toxisome and DON formation, was associated with the down-regulation of trichothecenes biosynthesis genes expression and many energy metabolism pathways as well as the inhibition of lipid droplets, the disturbed energy homeostasis and intracellular structures might ultimately inhibit fungal growth and DON production. In addition, limonene formulation enhanced the antifungal activity of triazole fungicides tebuconazole and mefentrifluconazole against F. graminearum, indicated limonene formulation has valuable potential as a bio-alternative fungicide and eco-friendly compound preparation for the effective management of F. graminearum and DON contamination in agriculture.

Fungicide Resistance: Progress in Understanding Mechanism, Monitoring, and Management.

Fungicide treatments are often essential for maintaining healthy crops and to achieve reliable and high-quality yields. However, continued use of fungicides with the same modes of action can lead to development of fungicide resistance, which has emerged in various plant pathogens and is a serious threat to effective crop protection. Exploration of resistance mechanisms is critical for resistance monitoring and management. This brief review summarizes advances during the past five decades in understanding the molecular resistance mechanisms of plant pathogenic fungi and oomycetes to major classes of fungicides, including benzimidazoles, myosin inhibitors, sterol demethylation inhibitors, quinone outside inhibitors, succinate dehydrogenase inhibitors, anilinopyrimidines, carboxylic acid amides, and oxysterol-binding protein homolog inhibitors. Based on known resistance mechanisms, PCR- and loop-mediated isothermal amplification-based approaches have been developed to allow high-throughput monitoring and early/rapid detection of emerging resistance. Classical principles in fungicide resistance management are also summarized, including using different modes of action of fungicides, limiting the number of applications of the chemicals with site-specific modes of action, and avoidance of their eradicant use. Future studies on novel strategies of disease management, including development of epigenetics- and RNA-based fungicides, will provide valuable knowledge for management of fungicide resistance.

SUMOylation regulates low-temperature survival and oxidative DNA damage tolerance in Botrytis cinerea.

SUMOylation as one of the protein post-translational modifications plays crucial roles in multiple biological processes of eukaryotic organisms. Botrytis cinerea is a devastating fungal pathogen and capable of infecting plant hosts at low temperature. However, the molecular mechanisms of low-temperature adaptation are largely unknown in fungi. Combining with biochemical methods and biological analyses, we report that SUMOylation regulates pathogen survival at low temperature and oxidative DNA damage response during infection in B. cinerea. The heat shock protein (Hsp70) BcSsb and E3 ubiquitin ligase BcRad18 were identified as substrates of SUMOylation; moreover, their SUMOylation both requires a single unique SUMO-interacting motif (SIM). SUMOylated BcSsb regulates ß-tubulin accumulation, thereby affecting the stability of microtubules and consequently mycelial growth at low temperature. On the contrary, SUMOylated BcRad18 modulates mono-ubiquitination of the sliding clamp protein proliferating cell nuclear antigen (PCNA), which is involved in response to oxidative DNA damage during infection. Our study uncovers the molecular mechanisms of SUMOylation-mediated low-temperature survival and oxidative DNA damage tolerance during infection in a devastating fungal pathogen, which provides novel insights into low-temperature adaptation and pathogenesis for postharvest pathogens as well as new targets for inhibitor invention in disease control.

SUMOylation regulates pre-mRNA splicing to overcome DNA damage in fungi.

Pathogenic fungi are subject to DNA damage stress derived from host immune responses during infection. Small ubiquitin-like modifier (SUMO) modification and precursor (pre)-mRNA splicing are both involved in DNA damage response (DDR). However, the mechanisms of how SUMOylation and splicing coordinated in DDR remain largely unknown. Combining with biochemical analysis, RNA-Seq method, and biological analysis, we report that SUMO pathway participates in DDR and virulence in Fusarium graminearum, a causal agent of Fusarium head blight of cereal crops world-wide. Interestingly, a key transcription factor FgSR is SUMOylated upon DNA damage stress. SUMOylation regulates FgSR nuclear-cytoplasmic partitioning and its phosphorylation by FgMec1, and promotes its interaction with chromatin remodeling complex SWI/SNF for activating the expression of DDR-related genes. Moreover, the SWI/SNF complex was found to further recruit splicing-related NineTeen Complex, subsequently modulates pre-mRNA splicing during DDR. Our findings reveal a novel function of SUMOylation in DDR by regulating a transcription factor to orchestrate gene expression and pre-mRNA splicing to overcome DNA damage during the infection of F. graminearum, which advances the understanding of the delicate regulation of DDR by SUMOylation in pathogenic fungi, and extends the knowledge of cooperation of SUMOylation and pre-mRNA splicing in DDR in eukaryotes.

The Drug H+ Antiporter FgQdr2 Is Essential for Multiple Drug Resistance, Ion Homeostasis, and Pathogenicity in Fusarium graminearum.

Increased emergence of drug resistance and DON pollution pose a severe problem in Fusarium head blight (FHB) control. While the H+ antiporter (DHA) family plays crucial roles in drug resistance, the characterization of DHA transporters has not been systematically studied in pathogenetic fungi. In this study, a systematic gene deletion analysis of all putative DHA transporter genes was carried out in Fusarium graminearum, and one DHA1 transporter FgQdr2 was found to be involved in multiple drug resistance, ion homeostasis, and virulence. Further exploration showed that FgQdr2 is mainly localized in the cell membrane; its expression under normal growth conditions is comparatively low, but sufficient for the regulation of drug efflux. Additionally, investigation of its physiological substrates demonstrated that FgQdr2 is essential for the transport of K+, Na+, Cu2+, and the regulation of the membrane proton gradient. For its roles in the FHB disease cycle, FgQdr2 is associated with fungal infection via regulating the biosynthesis of virulence factor deoxynivalenol (DON), the scavenging of the phytoalexin, as well as both asexual and sexual reproduction in F. graminearum. Overall, the results of this study reveal the crucial roles of FgQdr2 in multiple drug resistance, ion homeostasis, and pathogenicity, which advance the understanding of the DHA transporters in pathogenetic fungi.

Toxicity and action mechanisms of silver nanoparticles against the mycotoxin-producing fungus Fusarium graminearum.

Introduction: Fusarium graminearum is a most destructive fungal pathogen that causes Fusarium head blight (FHB) disease in cereal crops, resulting in severe yield loss and mycotoxin contamination in food and feed. Silver nanoparticles (AgNPs) are extensively applied in multiple fields due to their strong antimicrobial activity and are considered alternatives to fungicides. However, the antifungal mechanisms and the effects of AgNPs on mycotoxin production have not been well characterized. Objectives: This study aimed to investigate the antifungal activity and mechanisms of AgNPs against both fungicide-resistant and fungicide-sensitive F. graminearum strains, determine their effects on mycotoxin deoxynivalenol (DON) production, and evaluate the potential of AgNPs for FHB management in the field. Methods: Scanning electron microscopy (SEM), transmission electron microscopy (TEM), and fluorescence microscopy were used to examine the fungal morphological changes caused by AgNPs. In addition, RNA-Seq, qRT-PCR, and western blotting were conducted to detect gene transcription and DON levels. Results: AgNPs with a diameter of 2 nm exhibited effective antifungal activity against both fungicide-sensitive and fungicide-resistant strains of F. graminearum. Further studies showed that AgNP application could impair the development, cell structure, cellular energy utilization, and metabolism pathways of this fungus. RNA-Seq analysis and sensitivity determination revealed that AgNP treatment significantly induced the expression of azole-related ATP-binding cassette (ABC) transporters without compromising the control efficacy of azoles in F. graminearum. AgNP treatment stimulated the generation of reactive oxygen species (ROS), subsequently induced transcription of DON biosynthesis genes, toxisome formation, and mycotoxin production. Conclusion: This study revealed the underlying mechanisms of AgNPs against F. graminearum, determined their effects on DON production, and evaluated the potential of AgNPs for controlling fungicide-resistant F. graminearum strains. Together, our findings suggest that combinations of AgNPs with DON-reducing fungicides could be used for the management of FHB in the future.

The Importin FgPse1 Is Required for Vegetative Development, Virulence, and Deoxynivalenol Production by Interacting with the Nuclear Polyadenylated RNA-Binding Protein FgNab2 in Fusarium graminearum.

Karyopherins are involved in transport through nuclear pore complexes. Karyopherins are necessary for nuclear import and export pathways and bind to their cargos. Polyadenylation of messenger RNA (mRNA) is necessary for various biological processes, regulating gene expression in eukaryotes. Until now, the association of karyopherin with mRNA polyadenylation has been less understood in plant pathogenic fungi. In our study, we focused on the biological functions of the karyopherin FgPse1 in Fusarium graminearum. The results showed that FgPse1 is involved in mycelial growth, asexual reproduction, virulence, and deoxynivalenol (DON) production. Co-immunoprecipitation and bimolecular fluorescence complementation showed that FgPse1 interacts with the nuclear polyadenylated RNA-binding protein FgNab2. Moreover, a fluorescence localization assay indicated that FgPse1 is necessary for the nuclear import of FgNab2. The nuclear import of FgNab2 regulates the expression of FgTri4, FgTri5, and FgTri6, which are essential for DON production. Thus, ΔFgPse1 and ΔFgNab2 showed consistent defects in DON production. In summary, our data indicated that FgPse1 is necessary for mycelial growth, virulence, and DON production, interacting with FgNab2 in F. graminearum. These results contribute to our understanding of the functions of importins in phytopathogenic fungi.

Plant defense compound triggers mycotoxin synthesis by regulating H2B ub1 and H3K4 me2/3 deposition.

Fusarium graminearum produces the mycotoxin deoxynivalenol (DON) which promotes its expansion during infection on its plant host wheat. Conditional expression of DON production during infection is poorly characterized. Wheat produces the defense compound putrescine, which induces hypertranscription of DON biosynthetic genes (FgTRIs) and subsequently leads to DON accumulation during infection. Further, the regulatory mechanisms of FgTRIs hypertranscription upon putrescine treatment were investigated. The transcription factor FgAreA regulates putrescine-mediated transcription of FgTRIs by facilitating the enrichment of histone H2B monoubiquitination (H2B ub1) and histone 3 lysine 4 di- and trimethylations (H3K4 me2/3) on FgTRIs. Importantly, a DNA-binding domain (bZIP) specifically within the Fusarium H2B ub1 E3 ligase Bre1 othologs is identified, and the binding of this bZIP domain to FgTRIs depends on FgAreA-mediated chromatin rearrangement. Interestingly, H2B ub1 regulates H3K4 me2/3 via the methyltransferase complex COMPASS component FgBre2, which is different from Saccharomyces cerevisiae. Taken together, our findings reveal the molecular mechanisms by which host-generated putrescine induces DON production during F. graminearum infection. Our results also provide a novel insight into the role of putrescine during phytopathogen-host interactions and broaden our knowledge of H2B ub1 biogenesis and crosstalk between H2B ub1 and H3K4 me2/3 in eukaryotes.

Interplay of two transcription factors for recruitment of the chromatin remodeling complex modulates fungal nitrosative stress response.

Nitric oxide (NO) is a diffusible signaling molecule that modulates animal and plant immune responses. In addition, reactive nitrogen species derived from NO can display antimicrobial activities by reacting with microbial cellular components, leading to nitrosative stress (NS) in pathogens. Here, we identify FgAreB as a regulator of the NS response in Fusarium graminearum, a fungal pathogen of cereal crops. FgAreB serves as a pioneer transcription factor for recruitment of the chromatin-remodeling complex SWI/SNF at the promoters of genes involved in the NS response, thus promoting their transcription. FgAreB plays important roles in fungal infection and growth. Furthermore, we show that a transcription repressor (FgIxr1) competes with the SWI/SNF complex for FgAreB binding, and negatively regulates the NS response. NS, in turn, promotes the degradation of FgIxr1, thus enhancing the recruitment of the SWI/SNF complex by FgAreB.

The RasGEF FgCdc25 regulates fungal development and virulence in Fusarium graminearum via cAMP and MAPK signalling pathways.

Ras GTPases act as molecular switches to control various cellular processes by coupling integrated signals in eukaryotes. Activities of Ras GTPases are triggered by Ras GTPase guanine nucleotide exchange factors (RasGEFs) in general, whereas the role of RasGEF in plant pathogenic fungi is largely unknown. In this study, we characterized the only RasGEF protein in Fusarium graminearum, FgCdc25, by combining genetic, cytological and phenotypic strategies. FgCdc25 directly interacted with RasGTPase FgRas2, but not FgRas1, to regulate growth and sexual reproduction. Mutation of the FgCDC25 gene resulted in decreased toxisome formation and deoxynivalenol (DON) production, which was largely depended on cAMP signalling. In addition, FgCdc25 indirectly interacted with FgSte11 in FgSte11-Ste7-Gpmk1 cascade, and the ΔFgcdc25 strain totally abolished the formation of infection structures and was nonpathogenic in planta, which was partially recovered by addition of exogenous cAMP. In contrast, FgCdc25 directly interplayed with FgBck1 in FgBck1-MKK1-Mgv1 cascade to negatively control cell wall integrity. Collectively, these results suggest that FgCdc25 modulates cAMP and MAPK signalling pathways and further regulates fungal development, DON production and plant infection in F. graminearum.

Functional Analysis and Genome Mining Reveal High Potential of Biocontrol and Plant Growth Promotion in Nodule-Inhabiting Bacteria Within Paenibacillus polymyxa Complex.

Bacteria belonging to the genus Paenibacillus were frequently isolated from legume nodules. The nodule-inhabiting Paenibacillus as a resource of biocontrol and plant growth-promoting endophytes has rarely been explored. This study explored the nodule-inhabiting Paenibacillus' antifungal activities and biocontrol potentials against broad-spectrum important phytopathogenic fungi. We collected strains which were isolated from nodules of Robinia pseudoacacia, Dendrolobium triangulare, Ormosia semicastrata, Cicer arietinum, Acacia crassicarpa, or Acacia implexa and belong to P. peoriae, P. kribbensis, P. endophyticus, P. enshidis, P. puldeungensis, P. taichungensis, or closely related to P. kribbensis, or P. anseongense. These nodule-inhabiting Paenibacillus showed diverse antagonistic activities against five phytopathogenic fungi (Fusarium graminearum, Magnaporthe oryzae, Rhizoctonia solani, Sclerotinia sclerotiorum, and Botrytis cinerea). Six strains within the P. polymyxa complex showed broad-spectrum and potent activities against all the five pathogens, and produced multiple hydrolytic enzymes, siderophores, and lipopeptide fusaricidins. Fusaricidins are likely the key antimicrobials responsible for the broad-spectrum antifungal activities. The nodule-inhabiting strains within the P. polymyxa complex were able to epiphytically and endophytically colonize the non-host wheat plants, produce indole acetic acids (IAA), and dissolve calcium phosphate and calcium phytate. P. peoriae strains RP20, RP51, and RP62 could fix N2. P. peoriae RP51 and Paenibacillus sp. RP31, which showed potent plant colonization and plant growth-promotion competence, effectively control fungal infection in planta. Genome mining revealed that all strains (n = 76) within the P. polymyxa complex contain ipdC gene encoding indole-3-pyruvate decarboxylase for biosynthesis of IAA, 96% (n = 73) contain the fus cluster for biosynthesis of fusaricidins, and 43% (n = 33) contain the nif cluster for nitrogen fixation. Together, our study highlights that endophytic strains within the P. polymyxa complex have a high probability to be effective biocontrol agents and biofertilizers and we propose an effective approach to screen strains within the P. polymyxa complex.

A fungal ABC transporter FgAtm1 regulates iron homeostasis via the transcription factor cascade FgAreA-HapX.

Iron homeostasis is important for growth, reproduction and other metabolic processes in all eukaryotes. However, the functions of ATP-binding cassette (ABC) transporters in iron homeostasis are largely unknown. Here, we found that one ABC transporter (named FgAtm1) is involved in regulating iron homeostasis, by screening sensitivity to iron stress for 60 ABC transporter mutants of Fusarium graminearum, a devastating fungal pathogen of small grain cereal crops worldwide. The lack of FgAtm1 reduces the activity of cytosolic Fe-S proteins nitrite reductase and xanthine dehydrogenase, which causes high expression of FgHapX via activating transcription factor FgAreA. FgHapX represses transcription of genes for iron-consuming proteins directly but activates genes for iron acquisition proteins by suppressing another iron regulator FgSreA. In addition, the transcriptional activity of FgHapX is regulated by the monothiol glutaredoxin FgGrx4. Furthermore, the phosphorylation of FgHapX, mediated by the Ser/Thr kinase FgYak1, is required for its functions in iron homeostasis. Taken together, this study uncovers a novel regulatory mechanism of iron homeostasis mediated by an ABC transporter in an important pathogenic fungus.

A phosphorylated transcription factor regulates sterol biosynthesis in Fusarium graminearum.

Sterol biosynthesis is controlled by transcription factor SREBP in many eukaryotes. Here, we show that SREBP orthologs are not involved in the regulation of sterol biosynthesis in Fusarium graminearum, a fungal pathogen of cereal crops worldwide. Instead, sterol production is controlled in this organism by a different transcription factor, FgSR, that forms a homodimer and binds to a 16-bp cis-element of its target gene promoters containing two conserved CGAA repeat sequences. FgSR is phosphorylated by the MAP kinase FgHog1, and the phosphorylated FgSR interacts with the chromatin remodeling complex SWI/SNF at the target genes, leading to enhanced transcription. Interestingly, FgSR orthologs exist only in Sordariomycetes and Leotiomycetes fungi. Additionally, FgSR controls virulence mainly via modulating deoxynivalenol biosynthesis and responses to phytoalexin.

Lipid droplet biogenesis regulated by the FgNem1/Spo7-FgPah1 phosphatase cascade plays critical roles in fungal development and virulence in Fusarium graminearum.

Lipid droplets (LDs) control lipid metabolism in eukaryotic cells in general. However, the biogenesis regulation and biological functions of LDs are largely unknown in pathogenic fungi. Rapamycin treatment results in a significant increase of LD biogenesis in Fusarium graminearum. Molecular mechanisms of the target of rapamycin (TOR) pathway in regulating LD biogenesis and the functions of LD in virulence of F. graminearum were investigated in depth by combining genetic, cytological and phenotypic strategies. TOR in Fusarium graminearum (FgTOR) inhibition by rapamycin induces LD biogenesis through the FgPpg1/Sit4 signaling branch. FgPpg1 promotes phosphorylation of protein phosphatase FgNem1 by the protein kinase FgCak1. The phosphorylated FgNem1 dephosphorylates the phosphatidate phosphatase FgPah1. Dephosphorylated FgPah1 is active and stimulates LD biogenesis. Moreover, deletion of FgNem1/Spo7 or FgPah1 leads to serious defects in vegetative growth, sexual development and virulence. The results of this study provide novel insights into the regulatory mechanism and biological functions of the LDs in the devastating pathogenic fungus F. graminearum.

The MAPK kinase BcMkk1 suppresses oxalic acid biosynthesis via impeding phosphorylation of BcRim15 by BcSch9 in Botrytis cinerea.

The mitogen-activated protein kinase (MAPK) cassette of the cell wall integrity (CWI) pathway is primarily responsible for orchestrating changes of cell wall. However, functions of this cassette in other cellular processes are not well understood. Here, we found that the Botrytis cinerea mutant of MAPK kinase (BcMkk1) displays more serious defects in mycelial growth, conidiation, responses to cell wall and oxidative stresses, but possesses less reduced virulence than the mutants of its upstream (BcBck1) and downstream (BcBmp3) kinases. Interestingly, BcMkk1, but not BcBck1 and BcBmp3, negatively regulates production of oxalic acid (OA) and activity of extracellular hydrolases (EHs) that are proposed to be virulence factors of B. cinerea. Moreover, we obtained evidence that BcMkk1 negatively controls OA production via impeding phosphorylation of the Per-Arnt-Sim (PAS) kinase BcRim15 by the Ser/Thr kinase BcSch9. In addition, the fungal Pro40 homolog BcPro40 was found to interact simultaneously with three MAPKs, implying that BcPro40 is a scaffold protein of the CWI pathway in B. cinerea. Taken together, results of this study reveal that BcMkk1 negatively modulates virulence via suppressing OA biosynthesis in B. cinerea, which provides novel insight into conserved and species-specific functions of the MAPK kinase in fungi.

The ATP-binding protein FgArb1 is essential for penetration, infectious and normal growth of Fusarium graminearum.

ATP-binding cassette (ABC) transporters act mainly to transport compounds across cellular membranes and are important for diverse biological processes. However, their roles in pathogenesis have not been well-characterized in Fusarium graminearum. Sixty F. graminearum ABC protein genes were functionally characterized. Among them, FgArb1 regulates normal growth and importantly is essential for pathogenicity. Thus, the regulatory mechanisms of FgArb1 in pathogenicity were analyzed in this study. FgArb1 interacts with the mitogen-activated protein kinase (MAPK) FgSte7, and partially modulates plant penetration by regulating the phosphorylation of FgGpmk1 (the downstream kinase of FgSte7). The FgArb1 mutant exhibited dramatically reduced infective growth within wounded host tissues, likely resulting from its increased sensitivity to oxidative stresses, defective cell wall integrity and reduced deoxynivalenol (DON) production. FgArb1 also is important for the production of sexual and asexual spores that are important propagules for plant infection. In addition, FgArb1 is involved in the regulation of protein biosynthesis through impeding nuclear export of small ribosomal subunit. Finally, acetylation modification at sites K28, K65, K341 and K525 in FgArb1 is required for its biological functions. Taken together, results of this study provide a novel insight into functions of the ABC transporter in fungal pathogenesis.

The Activators of Type 2A Phosphatases (PP2A) Regulate Multiple Cellular Processes Via PP2A-Dependent and -Independent Mechanisms in Fusarium graminearum.

The type 2A protein phosphatases (PP2As) are holoenzymes in all eukaryotes but their activators remain unknown in filamentous fungi. Fusarium graminearum contains three PP2As (FgPp2A, FgSit4, and FgPpg1), which play critical roles in fungal growth, development, and virulence. Here, we identified two PP2A activators (PTPAs), FgRrd1 and FgRrd2, and found that they control PP2A activity in a PP2A-specific manner. FgRrd1 interacts with FgPpg1, but FgRrd2 interacts with FgPp2A and very weakly with FgSit4. Furthermore, FgRrd2 activates FgPp2A via regulating FgPp2A methylation. Phenotypic assays showed that FgRrd1 and FgRrd2 regulate mycelial growth, conidiation, sexual development, and lipid droplet biogenesis. More importantly, both FgRrd1 and FgRrd2 interact with RNA polymerase II, subsequently modulating its enrichments at the promoters of mycotoxin biosynthesis genes, which is independent on PP2A. In addition, FgRrd2 modulates response to phenylpyrrole fungicide, via regulating the phosphorylation of kinase FgHog1 in the high-osmolarity glycerol pathway, and to caffeine, via modulating FgPp2A methylation. Taken together, results of this study indicate that FgRrd1 and FgRrd2 regulate multiple physiological processes via different regulatory mechanisms in F. graminearum, which provides a novel insight into understanding the biological functions of PTPAs in fungi.

The FgSsb-FgZuo-FgSsz complex regulates multiple stress responses and mycotoxin production via folding the soluble SNARE Vam7 and ß2-tubulin in Fusarium graminearum.

Hsp70 proteins play important roles in protein folding in the budding yeast, but their functions in pathogenic fungi are largely unknown. Here, we found that Fusarium graminearum Hsp70 proteins FgSsb, FgSsz and their cochaperone FgZuo formed a complex. This complex was required for microtubule morphology, vacuole fusion and endocytosis. More importantly, the ß2-tubulin FgTub2 and SNARE protein FgVam7 were identified as targeting proteins of this complex. We further found that the complex FgSsb-FgZuo-FgSsz controlled sensitivity of F. graminearum to the antimicrotubule drug carbendazim and cold stress via regulating the folding of FgTub2. Moreover, this complex assisted the folding of FgVam7, subsequently modulated vacuole fusion and responses to heavy metal, osmotic and oxidative stresses. In addition, the deletion of this complex led to dramatically decreased deoxynivalenol biosynthesis. This study uncovers a novel regulating mechanism of Hsp70 in multiple stress responses in a filamentous fungus.