The G-protein alpha subunit AaGA1 positively regulates vegetative growth, appressorium-like formation, and pathogenicity in Alternaria alternata.

AIMS: The Gα subunit is a major component of heterotrimeric G proteins, which play a crucial role in the development and pathogenicity of several model fungi. However, its detailed function in the causal agent of pear black spot (Alternaria alternata) is unclear. Our aim was to understand the characteristics and functions of AaGA1 in A. alternata. METHODS AND RESULTS: AaGA1 was cloned from A. alternata in this study, which encodes 353 amino acids and has a "G-alpha" domain. Mutant ΔAaGA1 resulted in reduced vegetative growth, conidiation, and spore germination. Especially, mutant ΔAaGA1 produced only fewer conidia on the V8A medium, and spore formation-related genes AbaA, BrlA, and WetA were significantly downregulated. More tolerance against cell wall-inhibiting agents was observed after the deletion of AaGA1. Moreover, AaGA1 deletion led to a significant reduction in melanin and toxin production. Interestingly, deletion of AaGA1 resulted in defective appressorium-like formations, complete loss of the ability to penetrate cellophane, and decreased infection on non-wound inoculated tobacco leaves. Cell wall-degrading enzyme-related genes PME, CL, Cut2, and LC were significantly downregulated in mutant ΔAaGA1 mutant, significantly reducing virulence on wound-inoculated pear fruits. CONCLUSIONS: The G protein alpha subunit AaGA1 is indispensable for fungal development, appressorium-like formations, and pathogenicity in A. alternata.

Preharvest spraying of phenylalanine activates the sucrose and respiratory metabolism in muskmelon wounds during healing.

Phenylalanine (Phe) accelerates fruit wound healing by activating phenylpropanoid metabolism. However, whether Phe affects sucrose and respiratory metabolism in fruit during wound healing remains unknown. In this research, we found that preharvest Phe spray promoted sucrose degradation and increased glucose and fructose levels by activating acid invertase (AI), neutral invertase (NI), sucrose synthase (SS) and sucrose phosphate synthase (SPS) on harvested muskmelons. The spray also activated hexokinase (HK), phosphofructokinase (PFK), pyruvate kinase (PK), malate dehydrogenase (MDH), succinate dehydrogenase (SDH) and glucose-6-phosphate dehydrogenase (G6PDH). In addition, the spray improved energy and reducing power levels in the fruit. Taken together, preharvest Phe spray can provide carbon skeleton, energy and reducing power for wound healing by activating the sucrose metabolism, Embden-Meyerhof-Parnas (EMP) pathway, tricarboxylic acid (TCA) cycle and pentose phosphate (PPP) pathway in muskmelon wounds during healing, which is expected to be developed as a new strategy to accelerate fruit wound healing.

Essential role of ABA signaling and related transcription factors in phenolic acid and lignin synthesis during muskmelon wound healing.

Abscisic acid (ABA) is a key phytohormone involved in wound healing in fruits and vegetables, while fluridone (FLD) is its synthetic inhibitor. However, it is unknown whether ABA signaling and downstream transcription factors are involved in the synthesis of phenolic acids and lignin monomers in muskmelon wounds, and the underlying mechanisms. In our study, exogenous ABA promoted endogenous ABA synthesis by increasing the levels of ß-carotenoid and zeaxanthin, activating 9-cis-epoxycarotenoid dioxygenase (NCED) and zeaxanthin epoxidase (ZEP), facilitated ABA signaling by increasing the expression levels of protein phosphatases type 2C (CmPP2C) and ABA-responsive element binding factors (CmABF), upregulated the expression levels of CmMYB1 and CmWRKY1, and ABA induced phenylpropanoid metabolism by activating phenylalanine ammonia-lyase (PAL), 4-coenzyme A ligase (4CL), and cinnamyl alcohol dehydrogenase (CAD), which further increased the synthesis of phenolic acids and lignin monomers in muskmelon wounds during healing. Taken together, exogenous ABA induced phenylpropanoid metabolism and increased the synthesis of phenolic acid and lignin monomer in muskmelon wounds during healing, and may be involved in endogenous ABA synthesis and signaling and related transcription factors.

MYB24, MYB144, and MYB168 positively regulate suberin biosynthesis at potato tuber wounds during healing.

The essence of wound healing is the accumulation of suberin at wounds, which is formed by suberin polyphenolic (SPP) and suberin polyaliphatic (SPA). The biosynthesis of SPP and SPA monomers is catalyzed by several enzyme classes related to phenylpropanoid metabolism and fatty acid metabolism, respectively. However, how suberin biosynthesis is regulated at the transcriptional level during potato (Solanum tuberosum) tuber wound healing remains largely unknown. Here, 6 target genes and 15 transcription factors related to suberin biosynthesis in tuber wound healing were identified by RNA-seq technology and qRT-PCR. Dual luciferase and yeast one-hybrid assays showed that StMYB168 activated the target genes StPAL, StOMT, and St4CL in phenylpropanoid metabolism. Meanwhile, StMYB24 and StMYB144 activated the target genes StLTP, StLACS, and StCYP in fatty acid metabolism, and StFHT involved in the assembly of SPP and SPA domains in both native and wound periderms. More importantly, virus-induced gene silencing in S. tuberosum and transient overexpression in Nicotiana benthamiana assays confirmed that StMYB168 regulates the biosynthesis of free phenolic acids, such as ferulic acid. Furthermore, StMYB24/144 regulated the accumulation of suberin monomers, such as ferulates, α, ω-diacids, and ω-hydroxy acids. In conclusion, StMYB24, StMYB144, and StMYB168 have an elaborate division of labor in regulating the synthesis of suberin during tuber wound healing.

The sensor protein AaSho1 regulates infection structures differentiation, osmotic stress tolerance and virulence via MAPK module AaSte11-AaPbs2-AaHog1 in Alternaria alternata.

The high-osmolarity-sensitive protein Sho1 functions as a key membrane receptor in phytopathogenic fungi, which can sense and respond to external stimuli or stresses, and synergistically regulate diverse fungal biological processes through cellular signaling pathways. In this study, we investigated the biological functions of AaSho1 in Alternaria alternata, the causal agent of pear black spot. Targeted gene deletion revealed that AaSho1 is essential for infection structure differentiation, response to external stresses and synthesis of secondary metabolites. Compared to the wild-type (WT), the ∆AaSho1 mutant strain showed no significant difference in colony growth, morphology, conidial production and biomass accumulation. However, the mutant strain exhibited significantly reduced levels of melanin production, cellulase (CL) and ploygalacturonase (PG) activities, virulence, resistance to various exogenous stresses. Moreover, the appressorium and infection hyphae formation rates of the ∆AaSho1 mutant strain were significantly inhibited. RNA-Seq results showed that there were four branches including pheromone, cell wall stress, high osmolarity and starvation in the Mitogen-activated Protein Kinase (MAPK) cascade pathway. Furthermore, yeast two-hybrid experiments showed that AaSho1 activates the MAPK pathway via AaSte11-AaPbs2-AaHog1. These results suggest that AaSho1 of A. alternata is essential for fungal development, pathogenesis and osmotic stress response by activating the MAPK cascade pathway via Sho1-Ste11-Pbs2-Hog1.

Preharvest salicylic acid application improves the amino acid content and volatile profile in Vitis vinifera L. cv. Chardonnay during development.

Nitrogen is an important component that affects grapevine growth and the formation of flavor-associated volatile chemicals in grape berries. Dynamic changes in amino acids and aroma compounds in Chardonnay grape berry preharvest treated with different doses of salicylic acid (SA) at onset and one week later of veraison stage were evaluated. Exogenous 1- or 3-mM SA application significantly increased the content of total soluble solid and titratable acid in grapes, while 5 mM SA tended to decrease their levels. Compared with the control, the concentration of yeast assimilable nitrogen were 9.3% and 14.6% higher in 3 mM SA-treated grapes in 2021 and 2022, respectively. Preharvest 3 mM SA treatment efficiently enhanced the accumulation of nine amino acids, including tryptophan, phenylalanine, tyrosine, aspartic acid, lysine, asparagine, valine, isoleucine and histidine, as well as the concentration of total amino acid with and without proline in the two grape vintages. Higher concentrations of primary phenylalanine-derivatives and terpenoids and lower levels of C6 compounds in grapes treated with 3 mM SA were observed during the 2021-2022 season. Overall, SA improved the quality of wine grape in a dose dependent manner, while the response of berries to SA treatment also showed effects of the vintage.

Chitosan and chitooligosaccharide regulated reactive oxygen species homeostasis at wounds of pear fruit during healing.

Both chitosan (CTS) and chitooligosaccharide (COS) can promote fruit healing. However, whether the two chemicals regulate reactive oxygen species (ROS) homeostasis during wound healing of pear fruit remains unknown. In this study, the wounded pear fruit (Pyrus bretschneideri cv. Dongguo) was treated with a 1 g L-1 CTS and COS. We found CTS and COS treatments increased NADPH oxidase and superoxide dismutase activities, and promoted O2.- and H2O2 production at wounds. CTS and COS also enhanced the activities of catalase, peroxidase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, and glutathione reductase, and elevated the levels of ascorbic acid and glutathione. In addition, the two chemicals improved antioxidant capacity in vitro and maintained cell membrane integrity at fruit wounds during healing. Taken together, CTS and COS can regulate ROS homeostasis at wounds of pear fruit during healing by scavenging excessive H2O2 and improving antioxidant capacity. Overall, the COS demonstrated superior performance over the CTS.

Chitooligosaccharide accelerated wound healing in potato tubers by promoting the deposition of suberin polyphenols and lignin at wounds.

Chitooligosaccharide (COS) is a low molecular weight product of chitosan degradation. Although COS induces plant resistance by activating phenylpropanoid metabolism, there are few reports on whether COS accelerates wound healing in potato tubers by promoting the deposition of phenolic acids and lignin monomers at wounds. The results showed that COS activated phenylalanine ammonialyase and cinnamate 4-hydroxylase and promoted the synthesis of cinnamic, caffeic, p-coumaric, ferulic acids, total phenolics and flavonoids. COS activated 4-coumaric acid coenzyme A ligase and cinnamyl alcohol dehydrogenase and promoted the synthesis of sinapyl, coniferyl and cinnamyl alcohols. COS also increased H2O2 levels and peroxidase activity and accelerated the deposition of suberin polyphenols and lignin on wounds. In addition, COS reduced weight loss and inhibited lesion expansion in tubers inoculated with Fusarium sulfureum. Taken together, COS accelerated wound healing in potato tubers by inducing phenylpropanoid metabolism and accelerating the deposition of suberin polyphenols and lignin at wounds.

Sodium silicate treatment accelerates biosynthesis and polymerization of suberin polyaliphatics monomers at wounds of muskmelon.

Suberin polyaliphatics (SPA) is an important component of healing closing layer at fruit wounds. However, few study is available on the effect of sodium silicon treatment on SPA monomers biosynthesis and polymerization at muskmelon wounds. In this study, sodium silicate enhanced PLA2 (Phospholipase A2, PLA2) expression and enzyme activity, increased oleic acid, linoleic acid, and linolenic acid contents, and degree of fatty acids unsaturation at wounds. Sodium silicate upregulated the expressions of LACS4 (Long chain acyl CoA synthetase, LACS), KCS10 (ß-ketoacyl CoA synthase, KCS), CYP86B1 (Cytochrome P450 oxygenase, CYP), FAR3 (Fatty acyl CoA reductase, FAR), GPAT1 (Glycerol-3-phosphate acyltransferase, GPAT) and ABCG6 (ATP-binding cassette transporter), as well as their enzymes activities and ABC content. It is suggested that sodium silicate accelerates the deposition of SPA at muskmelon wounds by increasing the degree of fatty acids unsaturation, and promoting SPA monomers biosynthesis.

AaCaMKs Positively Regulate Development, Infection Structure Differentiation and Pathogenicity in Alternaria alternata, Causal Agent of Pear Black Spot.

Calcium/calmodulin-dependent protein kinase (CaMK), a key downstream target protein in the Ca2+ signaling pathway of eukaryotes, plays an important regulatory role in the growth, development and pathogenicity of plant fungi. Three AaCaMKs (AaCaMK1, AaCaMK2 and AaCaMK3) with conserved PKC_like superfamily domains, ATP binding sites and ACT sites have been cloned from Alternaria alternata, However, their regulatory mechanism in A. alternata remains unclear. In this study, the function of the AaCaMKs in the development, infection structure differentiation and pathogenicity of A. alternata was elucidated through targeted gene disruption. The single disruption of AaCaMKs had no impact on the vegetative growth and spore morphology but significantly influenced hyphae growth, sporulation, biomass accumulation and melanin biosynthesis. Further expression analysis revealed that the AaCaMKs were up-regulated during the infection structure differentiation of A. alternata on hydrophobic and pear wax substrates. In vitro and in vivo analysis further revealed that the deletion of a single AaCaMKs gene significantly reduced the A. alternata conidial germination, appressorium formation and infection hyphae formation. In addition, pharmacological analysis confirmed that the CaMK specific inhibitor, KN93, inhibited conidial germination and appressorium formation in A. alternata. Meanwhile, the AaCaMKs genes deficiency significantly reduced the A. alternata pathogenicity. These results demonstrate that AaCaMKs regulate the development, infection structure differentiation and pathogenicity of A. alternata and provide potential targets for new effective fungicides.

Aabrm1-mediated melanin synthesis is essential to growth and development, stress adaption, and pathogenicity in Alternaria alternata.

Scytalone dehydratase (brm1) is one of the key enzymes in 1, 8-dihydroxynaphthalene (DHN) melanin synthesis, which mediates melanin biosythesis and regulates cell biological process of plant fungi, but its function in Alternaria alternata, the causal agent of pear black spot, is unclear. Brm1 in A. alternata was cloned, identified, and named as Aabrm1. An Aabrm1-deletion mutant was generated and revealed that the deletion of Aabrm1 leads to a significant decrease in melanin production and forms orange colony smooth spores. In addition, the deletion of Aabrm1 gene impaired infection structure information and penetration. The external stress resistance of ΔAabrm1 was significantly weakened, and, in particular, it is very sensitive to oxidative stress, and the contents of H2O2 and O2.- in ΔAabrm1 were significantly increased. Virulence of ΔAabrm1 was reduced in non-wound-inoculated pear leaves but not changed in wound-inoculated pear fruit. These results indicated that Aabrm1-mediated melanin synthesis plays an important role in the pathogenicity of A. alternata.

Dynamic Change of Carbon and Nitrogen Sources in Colonized Apples by Penicillium expansum.

Penicillium expansum is a necrotrophic pathogen, which actively kills host cells and obtains nutrients from dead cells to achieve infection. However, few reports have elucidated the differential levels of carbon and nitrogen sources over increasing distances of the leading edge in fungal colonized fruit tissues during colonization. Our results showed that the highest consumption of sucrose and fructose, as well as the accumulation of glucose, were found in the decayed region of P. expansum-colonized 'Delicious' apple fruit compared with the healthy region at the leading edge and the healthy region 6 mm away from the leading edge. As nitrogen sources, the contents of methionine, glutamate, leucine, valine, isoleucine and serine were the lowest in the decayed region compared with the healthy regions during colonization. In addition, the titratable acidity, oxalic acid, citric acid, succinic acid and malic acid showed the highest accumulation in the decayed region compared with the healthy regions. P. expansum colonization induced the accumulation of saturated fatty acids in the decayed region, while the level of unsaturated fatty acids was the lowest. These changes were not observed in the healthy regions. These results indicated that P. expansum kills cells in advance of its colonization in order to obtain the nutrients of the apple tissue from the distal leading tissue of the colonized apple. It is understood that more carbon and nitrogen sources are required for fungal colonization, and a stronger defense response against colonization occurred in the fruit, causing the transit of nutrients from the distal tissue to the infected sites.

Updating Insights into the Regulatory Mechanisms of Calcineurin-Activated Transcription Factor Crz1 in Pathogenic Fungi.

Ca2+, as a second messenger in cells, enables organisms to adapt to different environmental stresses by rapidly sensing and responding to external stimuli. In recent years, the Ca2+ mediated calcium signaling pathway has been studied systematically in various mammals and fungi, indicating that the pathway is conserved among organisms. The pathway consists mainly of complex Ca2+ channel proteins, calcium pumps, Ca2+ transporters and many related proteins. Crz1, a transcription factor downstream of the calcium signaling pathway, participates in regulating cell survival, ion homeostasis, infection structure development, cell wall integrity and virulence. This review briefly summarizes the Ca2+ mediated calcium signaling pathway and regulatory roles in plant pathogenic fungi. Based on discussing the structure and localization of transcription factor Crz1, we focus on the regulatory role of Crz1 on growth and development, stress response, pathogenicity of pathogenic fungi and its regulatory mechanisms. Furthermore, we explore the cross-talk between Crz1 and other signaling pathways. Combined with the important role and pathogenic mechanism of Crz1 in fungi, the new strategies in which Crz1 may be used as a target to explore disease control in practice are also discussed.

Chitooligosaccharide Maintained Cell Membrane Integrity by Regulating Reactive Oxygen Species Homeostasis at Wounds of Potato Tubers during Healing.

Chitooligosaccharide (COS) is a degradation product of chitosan. Although COS increased fruit resistance by regulating the metabolism of reactive oxygen species (ROS), few reports are available on whether COS regulates ROS homeostasis at wounds of potato tubers during healing. In this study, COS increased gene expression and activities of NADPH oxidase and superoxide dismutase, and promoted the generation of O2●- and H2O2. Moreover, COS increased gene expression and activities of catalase, peroxidase, and AsA-GSH cycle-related enzymes, as well as the levels of ascorbic acid and glutathione levels. In addition, COS elevated the scavenging ability of DPPH, ABTS+, and FRAP, and reduced cell membrane permeability and malondialdehyde content. Taken together, COS could maintain cell membrane integrity by eliminating excessive H2O2 and improving the antioxidant capacity in vitro, which contributes to the maintainance of cell membrane integrity at wounds of potato tubers during healing.

Transcription factor AacmrA mediated melanin synthesis regulates the growth, appressorium formation, stress response and pathogenicity of pear fungal Alternaria alternata.

CmrA, as transcription factor for regulating DHN-melanin synthesis, controls melanin synthesis gene expression, and also regulate growth, development, stress response and virulence of plant fungi. However, little is known about the roles of CmrA on infection structure formation, penetration and pathogenicity of pear fungal Alternaria alternata. Here, we identified cmrA gene in A. alternata and assigned as AacmrA, sequence alignment and phylogenetic analysis revealed that AacmrA is highly conserved among fungi and encoded protein contain two Cys2His2 zinc finger motifs and one Zn(II)2Cys6 zinc cluster protein motif. ΔAacmrA severely decreased melanin production and DHN melanin synthesis related genes expression. Deletion of AacmrA impaired the morphology of spore and hyphae. Spore germination and appressorium formation induced by hydrophobicity surfaces and fruit wax significantly decreased in ΔAacmrA mutant. ΔAacmrA mutants were more sensitive than the wild type to osmotic stress and cell wall inhibitors, especially more sensitive to oxidative stress. In addition, lesion diameter of pear fruit wound inoculated with the ΔAacmrA mutant was reduced by 40.8% compared with the wild type 12 d after inoculation. All findings of this study suggested that AacmrA is required for melanin biosynthesis, infection structure formation, and pathogenicity in A. alternata.

AaCaM is required for infection structure differentiation and secondary metabolites in pear fungal pathogen Alternaria alternata.

AIMS: Calmodulin (CaM), acts as a kind of multifunctional Ca2+ sensing protein, which is ubiquitous in fungi, is highly conserved across eukaryotes and is involved in the regulation of a range of physiological processes, including morphogenesis, reproduction and secondary metabolites biosynthesis. Our aim was to understand the characteristics and functions of AaCaM in Alternaria alternata, the causal agent of pear black spot. METHODS AND RESULTS: A 450 bp cDNA sequence of AaCaM gene of A. alternata was cloned by the PCR homology method. Sequence analysis showed that this protein encoded by AaCaM was a stable hydrophilic protein and had a high similarity to Neurospora crassa (CAA50271.1) and other fungi. RT-qPCR analysis determined that AaCaM was differentially upregulated during infection structural differentiation of A. alternata both on hydrophobic and pear wax extract-coated surface, with a 3.37-fold upregulation during the hydrophobic induced appressorium formation period (6 h) and a 1.46-fold upregulation during the infection hyphae formation period (8 h) following pear wax induction. Pharmaceutical analysis showed that the CaM-specific inhibitor, trifluoperazine (TFP), inhibited spore germination and appressorium formation, and affected toxins and melanin biosynthesis in A. alternata. CONCLUSIONS: AaCaM plays an important role in regulating infection structure differentiation and secondary metabolism of A. alternata. SIGNIFICANCE AND IMPACT OF STUDY: Our study provides a theoretical basis for further in-depth investigation of the specific role of AaCaM in the calcium signalling pathway underlying hydrophobic and pear wax-induced infection structure differentiation and pathogenicity of A. alternata.

Molecular Characterization of Phospholipase C in Infection Structure Differentiation Induced by Pear Fruit Surface Signals, Stress Responses, Secondary Metabolism, and Virulence of Alternaria alternata.

Fungal pathogens use plant surface physiochemical signals to trigger specific developmental processes. To assess the role of phospholipase C (PLC) in mediating plant stimuli sensing of Alternaria alternata, the function of three PLC genes was characterized by constructing ΔAaPLC mutants. Here we showed that fruit wax-coated surfaces significantly induced appressorium formation in A. alternata and mutants. Germination of ΔAaPLC mutants did not differ from the wild type. Deletion of AaPLC1 led to the decrease of appressorium formation and infected hyphae, but the degree of reduction varies between the different types of waxes, with the strongest response to pear wax. Appressorium formation and infected hyphae of the ΔAaPLC1 mutant on dewaxed onion epidermis mounted with pear wax (θ4) were reduced by 14.5 and 65.7% after 8 h incubation, while ΔAaPLC2 and ΔAaPLC3 formed the same infection hyphae as wild type. In addition, AaPLC1 mutation caused pleiotropic effects on fungal biological function, including growth deficiency, changes in stress tolerance, weakening of pathogenicity to the host, as well as destruction of mycotoxin synthesis. Both AaPLC2 and AaPLC3 genes were found to have some effects on stress response and mycotoxin production. Taken together, AaPLC genes differentially regulate the growth, stress response, pathogenicity, and secondary metabolism of A. alternata.

Brassinosteroid Accelerates Wound Healing of Potato Tubers by Activation of Reactive Oxygen Metabolism and Phenylpropanoid Metabolism.

Wound healing could effectively reduce the decay rate of potato tubers after harvest, but it took a long time to form typical and complete healing structures. Brassinosteroid (BR), as a sterol hormone, is important for enhancing plant resistance to abiotic and biotic stresses. However, it has not been reported that if BR affects wound healing of potato tubers. In the present study, we observed that BR played a positive role in the accumulation of lignin and suberin polyphenolic (SPP) at the wounds, and effectively reduced the weight loss and disease index of potato tubers (cv. Atlantic) during healing. At the end of healing, the weight loss and disease index of BR group was 30.8% and 23.1% lower than the control, respectively. Furthermore, BR activated the expression of StPAL, St4CL, StCAD genes and related enzyme activities in phenylpropanoid metabolism, and promoted the synthesis of lignin precursors and phenolic acids at the wound site, mainly by inducing the synthesis of caffeic acid, sinapic acid and cinnamyl alcohol. Meanwhile, the expression of StNOX was induced and the production of O2- and H2O2 was promoted, which mediated oxidative crosslinking of above phenolic acids and lignin precursors to form SPP and lignin. In addition, the expression level of StPOD was partially increased. In contrast, the inhibitor brassinazole inhibited phenylpropanoid metabolism and reactive oxygen metabolism, and demonstrated the function of BR hormone in healing in reverse. Taken together, the activation of reactive oxygen metabolism and phenylpropanoid metabolism by BR could accelerate the wound healing of potato tubers.

AaHog1 Regulates Infective Structural Differentiation Mediated by Physicochemical Signals from Pear Fruit Cuticular Wax, Stress Response, and Alternaria alternata Pathogenicity.

The high-osmolarity glycerol response kinase, Hog1, affects several cellular responses, but the precise regulatory role of the Hog1 mitogen-activated protein (MAP) kinase in the differentiation of the infective structure of Alternariaalternata induced by pear cuticular wax and hydrophobicity has not yet clarified. In this study, the AaHog1 in A. alternata was identified and functionally characterized. AaHog1 has threonine-glycine-tyrosine (TGY) phosphorylation sites. Moreover, the expression level of AaHog1 was significantly upregulated during the stages of appressorium formation of A. alternata on the fruit-wax-extract-coated GelBond hydrophobic film surface. Importantly, our results showed that the appressorium and infection hyphae formation rates were significantly reduced in ΔAaHog1 mutants. Furthermore, AaHog1 is beneficial for the growth and development, stress tolerance, virulence, and cell-wall-degrading enzyme activity of A. alternata. These findings may be useful for dissecting the AaHog1 regulatory mechanism in relation to the pathogenesis of A. alternata.

Cellular Responses Required for Oxidative Stress Tolerance of the Necrotrophic Fungus Alternaria alternata, Causal Agent of Pear Black Spot.

To establish successful infections in host plants, pathogenic fungi must sense and respond to an array of stresses, such as oxidative stress. In this study, we systematically analyzed the effects of 30 mM H2O2 treatment on reactive oxygen species (ROS) metabolism in Alternaria alternata. Results showed that 30 mM H2O2 treatment lead to increased O2- generation rate and H2O2 content, and simultaneously, increased the activities and transcript levels of NADPH oxidase (NOX). The activities and gene expression levels of enzymes related with ascorbic acid-glutathione cycle (AsA-GSH cycle) and thioredoxin systems, including superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), ascorbate peroxidase (AXP) and thioredoxin (TrxR), were remarkably enhanced by 30 mM H2O2 stress treatment. Additionally, 30 mM H2O2 treatment decreased the glutathione (GSH) content, whereas it increased the amount of oxidized glutathione (GSSG), dehydroascorbate (DHA) and ascorbic acid (AsA). These results revealed that cellular responses are required for oxidative stress tolerance of the necrotrophic fungus A. alternata.