scholarly journals The Autophagy Gene BcATG8 Regulates the Vegetative Differentiation and Pathogenicity of Botrytis cinerea

2018 ◽  
Vol 84 (11) ◽  
Author(s):  
Weichao Ren ◽  
Na Liu ◽  
Chengwei Sang ◽  
Dongya Shi ◽  
Mingguo Zhou ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Plant Pathogen ◽  
Fluorescent Protein ◽  
Degradation Process ◽  
Gray Mold ◽  
Broad Host Range ◽  
Physical Interaction ◽  
Content Type ◽  
Autophagy Gene ◽  
Mold Fungus

ABSTRACT Autophagy is a conserved degradation process that maintains intracellular homeostasis to ensure normal cell differentiation and development in eukaryotes. ATG8 is one of the key molecular components of the autophagy pathway. In this study, we identified and characterized BcATG8 , a homologue of Saccharomyces cerevisiae (yeast) ATG8 in the necrotrophic plant pathogen Botrytis cinerea . Yeast complementation experiments demonstrated that BcATG8 can functionally complement the defects of the yeast ATG8 null mutant. Direct physical interaction between BcAtg8 and BcAtg4 was detected in the yeast two-hybrid system. Subcellular localization assays showed that green fluorescent protein-tagged BcAtg8 (GFP-BcAtg8) localized in the cytoplasm as preautophagosomal structures (PAS) under general conditions but mainly accumulated in the lumen of vacuoles in the case of autophagy induction. Deletion of BcATG8 (Δ BcAtg8 mutant) blocked autophagy and significantly impaired mycelial growth, conidiation, sclerotial formation, and virulence. In addition, the conidia of the Δ BcAtg8 mutant contained fewer lipid droplets (LDs), and quantitative real-time PCR (qRT-PCR) assays revealed that the basal expression levels of the LD metabolism-related genes in the mutant were significantly different from those in the wild-type (WT) strain. All of these phenotypic defects were restored by gene complementation. These results indicate that BcATG8 is essential for autophagy to regulate fungal development, pathogenesis, and lipid metabolism in B. cinerea . IMPORTANCE The gray mold fungus Botrytis cinerea is an economically important plant pathogen with a broad host range. Although there are fungicides for its control, many classes of fungicides have failed due to its genetic plasticity. Exploring the fundamental biology of B. cinerea can provide the theoretical basis for sustainable and long-term disease management. Autophagy is an intracellular process for degradation and recycling of cytosolic materials in eukaryotes and is now known to be vital for fungal life. Here, we report studies of the biological role of the autophagy gene BcATG8 in B. cinerea . The results suggest that autophagy plays a crucial role in vegetative differentiation and virulence of B. cinerea .

scholarly journals Living Colors in the Gray Mold Pathogen Botrytis cinerea: Codon-Optimized Genes Encoding Green Fluorescent Protein and mCherry, Which Exhibit Bright Fluorescence

2011 ◽  
Vol 77 (9) ◽  
pp. 2887-2897 ◽  
Author(s):  
Michaela Leroch ◽  
Dennis Mernke ◽  
Dieter Koppenhoefer ◽  
Prisca Schneider ◽  
Andreas Mosbach ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Botrytis Cinerea ◽  
Fluorescent Protein ◽  
Gc Content ◽  
Gray Mold ◽  
Mrna Levels ◽  
Content Type ◽  
Mold Fungus ◽  
Encoding Gene ◽  
Green Fluorescent

ABSTRACTThe green fluorescent protein (GFP) and its variants have been widely used in modern biology as reporters that allow a variety of live-cell imaging techniques. So far, GFP has rarely been used in the gray mold fungusBotrytis cinereabecause of low fluorescence intensity. The codon usage ofB. cinereagenes strongly deviates from that of commonly used GFP-encoding genes and reveals a lower GC content than other fungi. In this study, we report the development and use of a codon-optimized version of theB. cinereaenhanced GFP (eGFP)-encoding gene (Bcgfp) for improved expression inB. cinerea. Both the codon optimization and, to a smaller extent, the insertion of an intron resulted in higher mRNA levels and increased fluorescence. Bcgfpwas used for localization of nuclei in germinating spores and for visualizing host penetration. We further demonstrate the use of promoter-Bcgfpfusions for quantitative evaluation of various toxic compounds as inducers of theatrBgene encoding an ABC-type drug efflux transporter ofB. cinerea. In addition, a codon-optimized mCherry-encoding gene was constructed which yielded bright red fluorescence inB. cinerea.

scholarly journals Novel Hypovirulence-Associated RNA Mycovirus in the Plant-Pathogenic Fungus Botrytis cinerea: Molecular and Biological Characterization

2015 ◽  
Vol 81 (7) ◽  
pp. 2299-2310 ◽  
Author(s):  
Lin Yu ◽  
Wen Sang ◽  
Ming-De Wu ◽  
Jing Zhang ◽  
Long Yang ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Fluorescent Protein ◽  
Rna Virus ◽  
Pathogenic Fungus ◽  
Ornamental Plants ◽  
Gray Mold ◽  
Open Reading Frames ◽  
Content Type ◽  
Molecular Features ◽  
Green Fluorescent

ABSTRACTBotrytis cinereais a pathogenic fungus causing gray mold on numerous economically important crops and ornamental plants. This study was conducted to characterize the biological and molecular features of a novel RNA mycovirus, Botrytis cinerea RNA virus 1 (BcRV1), in the hypovirulent strain BerBc-1 ofB. cinerea. The genome of BcRV1 is 8,952 bp long with two putative overlapped open reading frames (ORFs), ORF1 and ORF2, coding for a hypothetical polypeptide (P1) and RNA-dependent RNA polymerase (RdRp), respectively. A −1 frameshifting region (designated the KNOT element) containing a shifty heptamer, a heptanucleotide spacer, and an H-type pseudoknot was predicted in the junction region of ORF1 and ORF2. The −1 frameshifting role of the KNOT element was experimentally confirmed through determination of the production of the fusion protein red fluorescent protein (RFP)-green fluorescent protein (GFP) by the plasmid containing the constructdsRed-KNOT-eGFPinEscherichia coli. BcRV1 belongs to a taxonomically unassigned double-stranded RNA (dsRNA) mycovirus group. It is closely related to grapevine-associated totivirus 2 and Sclerotinia sclerotiorum nonsegmented virus L. BcRV1 in strain BerBc-1 was found capable of being transmitted vertically through macroconidia and horizontally to otherB. cinereastrains through hyphal contact. The presence of BcRV1 was found to be positively correlated with hypovirulence inB. cinerea, with the attenuation effects of BcRV1 on mycelial growth and pathogenicity being greatly affected by the accumulation level of BcRV1.

scholarly journals Gray Mold Populations in German Strawberry Fields Are Resistant to Multiple Fungicides and Dominated by a Novel Clade Closely Related to Botrytis cinerea

2012 ◽  
Vol 79 (1) ◽  
pp. 159-167 ◽  
Author(s):  
Michaela Leroch ◽  
Cecilia Plesken ◽  
Roland W. S. Weber ◽  
Frank Kauff ◽  
Gabriel Scalliet ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Host Preference ◽  
Gray Mold ◽  
Vegetable Production ◽  
Multiple Gene ◽  
Content Type ◽  
Chemical Treatments ◽  
Mold Fungus ◽  
Encoding Gene ◽  
Specific Species

ABSTRACT The gray mold fungus Botrytis cinerea is a major threat to fruit and vegetable production. Strawberry fields usually receive several fungicide treatments against Botrytis per season. Gray mold isolates from several German strawberry-growing regions were analyzed to determine their sensitivity against botryticides. Fungicide resistance was commonly observed, with many isolates possessing resistance to multiple (up to six) fungicides. A stronger variant of the previously described multidrug resistance (MDR) phenotype MDR1, called MDR1h, was found to be widely distributed, conferring increased partial resistance to two important botryticides, cyprodinil and fludioxonil. A 3-bp deletion mutation in a transcription factor-encoding gene, mrr1 , was found to be correlated with MDR1h. All MDR1h isolates and the majority of isolates with resistance to multiple fungicides were found to be genetically distinct. Multiple-gene sequencing confirmed that they belong to a novel clade, called Botrytis group S, which is closely related to B. cinerea and the host-specific species B. fabae . Isolates of Botrytis group S genotypes were found to be widespread in all German strawberry-growing regions but almost absent from vineyards. Our data indicate a clear subdivision of gray mold populations, which are differentially distributed according to their host preference and adaptation to chemical treatments.

scholarly journals The Two Cryptochrome/Photolyase Family Proteins Fulfill Distinct Roles in DNA Photorepair and Regulation of Conidiation in the Gray Mold Fungus Botrytis cinerea

2017 ◽  
Vol 83 (17) ◽  
Author(s):  
Kim C. Cohrs ◽  
Julia Schumacher
Keyword(s):  
Botrytis Cinerea ◽  
Molecular Mechanisms ◽  
Uv Light ◽  
Gray Mold ◽  
Pyrimidine Dimer ◽  
Constant Darkness ◽  
Dependent Manner ◽  
Content Type ◽  
Mold Fungus ◽  
Signaling Function

ABSTRACT The plant-pathogenic leotiomycete Botrytis cinerea is known for the strict regulation of its asexual differentiation programs by environmental light conditions. Sclerotia are formed in constant darkness; black/near-UV (NUV) light induces conidiation; and blue light represses both differentiation programs. Sensing of black/NUV light is attributed to proteins of the cryptochrome/photolyase family (CPF). To elucidate the molecular basis of the photoinduction of conidiation, we functionally characterized the two CPF proteins encoded in the genome of B. cinerea as putative positive-acting components. B. cinerea CRY1 (BcCRY1), a cyclobutane pyrimidine dimer (CPD) photolyase, acts as the major enzyme of light-driven DNA repair (photoreactivation) and has no obvious role in signaling. In contrast, BcCRY2, belonging to the cry-DASH proteins, is dispensable for photorepair but performs regulatory functions by repressing conidiation in white and especially black/NUV light. The transcription of bccry1 and bccry2 is induced by light in a White Collar complex (WCC)-dependent manner, but neither light nor the WCC is essential for the repression of conidiation through BcCRY2 when bccry2 is constitutively expressed. Further, BcCRY2 affects the transcript levels of both WCC-induced and WCC-repressed genes, suggesting a signaling function downstream of the WCC. Since both CPF proteins are dispensable for photoinduction by black/NUV light, the origin of this effect remains elusive and may be connected to a yet unknown UV-light-responsive system. IMPORTANCE Botrytis cinerea is an economically important plant pathogen that causes gray mold diseases in a wide variety of plant species, including high-value crops and ornamental flowers. The spread of disease in the field relies on the formation of conidia, a process that is regulated by different light qualities. While this feature has been known for a long time, we are just starting to understand the underlying molecular mechanisms. Conidiation in B. cinerea is induced by black/near-UV light, whose sensing is attributed to the action of cryptochrome/photolyase family (CPF) proteins. Here we report on the distinct functions of two CPF proteins in the photoresponse of B. cinerea. While BcCRY1 acts as the major photolyase in photoprotection, BcCRY2 acts as a cryptochrome with a signaling function in regulating photomorphogenesis (repression of conidiation).

scholarly journals Cyclophilin BcCyp2 Regulates Infection-Related Development to Facilitate Virulence of the Gray Mold Fungus Botrytis cinerea

2021 ◽  
Vol 22 (4) ◽  
pp. 1694
Author(s):  
Jiao Sun ◽  
Chen-Hao Sun ◽  
Hao-Wu Chang ◽  
Song Yang ◽  
Yue Liu ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Cyclic Adenosine Monophosphate ◽  
Adenosine Monophosphate ◽  
Gray Mold ◽  
Gene Encoding ◽  
Histone 3 ◽  
Mold Fungus ◽  
Related Development ◽  
Hyphal Morphology ◽  
Cellular Components

Cyclophilin (Cyp) and Ca2+/calcineurin proteins are cellular components related to fungal morphogenesis and virulence; however, their roles in mediating the pathogenesis of Botrytis cinerea, the causative agent of gray mold on over 1000 plant species, remain largely unexplored. Here, we show that disruption of cyclophilin gene BcCYP2 did not impair the pathogen mycelial growth, osmotic and oxidative stress adaptation as well as cell wall integrity, but delayed conidial germination and germling development, altered conidial and sclerotial morphology, reduced infection cushion (IC) formation, sclerotial production and virulence. Exogenous cyclic adenosine monophosphate (cAMP) rescued the deficiency of IC formation of the ∆Bccyp2 mutants, and exogenous cyclosporine A (CsA), an inhibitor targeting cyclophilins, altered hyphal morphology and prevented host-cell penetration in the BcCYP2 harboring strains. Moreover, calcineurin-dependent (CND) genes are differentially expressed in strains losing BcCYP2 in the presence of CsA, suggesting that BcCyp2 functions in the upstream of cAMP- and Ca2+/calcineurin-dependent signaling pathways. Interestingly, during IC formation, expression of BcCYP2 is downregulated in a mutant losing BcJAR1, a gene encoding histone 3 lysine 4 (H3K4) demethylase that regulates fungal development and pathogenesis, in B. cinerea, implying that BcCyp2 functions under the control of BcJar1. Collectively, our findings provide new insights into cyclophilins mediating the pathogenesis of B. cinerea and potential targets for drug intervention for fungal diseases.

scholarly journals Elevated Genetic Variation Within Virulence-Associated Botrytis cinerea Polygalacturonase Loci

2007 ◽  
Vol 20 (9) ◽  
pp. 1126-1137 ◽  
Author(s):  
Heather C. Rowe ◽  
Daniel J. Kliebenstein
Keyword(s):  
Genetic Variation ◽  
Botrytis Cinerea ◽  
Sequence Variation ◽  
Gray Mold ◽  
Published Data ◽  
Broad Host Range ◽  
Conserved Sequence ◽  
Evolutionary Forces ◽  
Genetic Subdivision ◽  
Distinct Cell

Botrytis cinerea, or gray mold, is a necrotrophic fungal pathogen of hundreds of plant species. The genetic diversity of B. cinerea may contribute to its broad host range; however, the level and structure of genetic variation at pathogenesis-associated loci has not been described. B. cinerea possesses six distinct cell-wall-degrading polygalacturonases (PGs), enzymes of demonstrated importance to pathogenesis and interaction with host plant defenses. Sequencing a collection of 34 B. cinerea isolates at three PG-encoding loci, BcPG1, BcPG2, and BcPG3, revealed limited evidence of host-mediated genetic subdivision within loci, yet suggested differences in the action of evolutionary forces among loci. BcPG1 and BcPG2 are highly polymorphic, particularly when compared with previously published data from nonpathogenicity loci, whereas BcPG3 is relatively conserved. Sequence variation at BcPG1 and BcPG2 did not appear to be associated with virulence on Arabidopsis leaves; however, BcPG2 variation showed a statistically significant association with growth rate on pectin. Rather than providing evidence for host-mediated genetic subdivision at individual PG loci, our data support specialization among PGs and the potential diversification of PGs interacting directly with host defenses.

Streptomyces botrytidirepellens sp. nov., a novel actinomycete with antifungal activity against Botrytis cinerea

2021 ◽  
Vol 71 (9) ◽  
Author(s):  
Mengqi Jiang ◽  
Xi Xu ◽  
Jia Song ◽  
Dongmei Li ◽  
Liyuan Han ◽  
...  
Keyword(s):  
Antifungal Activity ◽  
Botrytis Cinerea ◽  
Gene Clusters ◽  
Gray Mold ◽  
Phytopathogenic Fungi ◽  
Antagonistic Effect ◽  
Economic Losses ◽  
Content Type ◽  
Desferrioxamine B ◽  
Cell Free Filtrate

The fungal pathogen Botrytis cinerea is the causal agent of devastating gray mold diseases in many economically important fruits, vegetables, and flowers, leading to serious economic losses worldwide. In this study, a novel actinomycete NEAU-LD23T exhibiting antifungal activity against B. cinerea was isolated, and its taxonomic position was evaluated using a polyphasic approach. Based on the genotypic, phenotypic and chemotaxonomic data, it is concluded that the strain represents a novel species within the genus Streptomyces , for which the name Streptomyces botrytidirepellens sp. nov. is proposed. The type strain is NEAU-LD23T (=CCTCC AA 2019029T=DSM 109824T). In addition, strain NEAU-LD23T showed a strong antagonistic effect against B. cinerea (82.6±2.5%) and varying degrees of inhibition on nine other phytopathogenic fungi. Both cell-free filtrate and methanol extract of mycelia of strain NEAU-LD23T significantly inhibited mycelial growth of B. cinerea. To preliminarily explore the antifungal mechanisms, the genome of strain NEAU-LD23T was sequenced and analyzed. AntiSMASH analysis led to the identification of several gene clusters responsible for the biosynthesis of bioactive secondary metabolites with antifungal activity, including 9-methylstreptimidone, echosides, anisomycin, coelichelin and desferrioxamine B. Overall, this research provided us an excellent strain with considerable potential to use for biological control of tomato gray mold.

scholarly journals Regulation of Pathogenic Spore Germination by CgRac1 in the Fungal Plant Pathogen Colletotrichum gloeosporioides

2011 ◽  
Vol 10 (8) ◽  
pp. 1122-1130 ◽  
Author(s):  
Iris Nesher ◽  
Anna Minz ◽  
Leonie Kokkelink ◽  
Paul Tudzynski ◽  
Amir Sharon
Keyword(s):  
Cell Division ◽  
Plant Pathogen ◽  
Germ Tube ◽  
Colletotrichum Gloeosporioides ◽  
Fluorescent Protein ◽  
Nuclear Division ◽  
Simultaneous Formation ◽  
Content Type ◽  
Germ Tubes

ABSTRACT Colletotrichum gloeosporioides is a facultative plant pathogen: it can live as a saprophyte on dead organic matter or as a pathogen on a host plant. Different patterns of conidial germination have been recognized under saprophytic and pathogenic conditions, which also determine later development. Here we describe the role of CgRac1 in regulating pathogenic germination. The hallmark of pathogenic germination is unilateral formation of a single germ tube following the first cell division. However, transgenic strains expressing a constitutively active CgRac1 (CA-CgRac1) displayed simultaneous formation of two germ tubes, with nuclei continuing to divide in both cells after the first cell division. CA-CgRac1 also caused various other abnormalities, including difficulties in establishing and maintaining cell polarity, reduced conidial and hyphal adhesion, and formation of immature appressoria. Consequently, CA-CgRac1 isolates were completely nonpathogenic. Localization studies with cyan fluorescent protein (CFP)-CgRac1 fusion protein showed that the CgRac1 protein is abundant in conidia and in hyphal tips. Although the CFP signal was equally distributed in both cells of a germinating conidium, reactive oxygen species accumulated only in the cell that produced a germ tube, indicating that CgRac1 was active only in the germinating cell. Collectively, our results show that CgRac1 is a major regulator of asymmetric development and that it is involved in the regulation of both morphogenesis and nuclear division. Modification of CgRac1 activity disrupts the morphogenetic program and prevents fungal infection.

scholarly journals Analysis of the Molecular Dialogue Between Gray Mold (Botrytis cinerea) and Grapevine (Vitis vinifera) Reveals a Clear Shift in Defense Mechanisms During Berry Ripening

2015 ◽  
Vol 28 (11) ◽  
pp. 1167-1180 ◽  
Author(s):  
Jani Kelloniemi ◽  
Sophie Trouvelot ◽  
Marie-Claire Héloir ◽  
Adeline Simon ◽  
Bérengère Dalmais ◽  
...  
Keyword(s):  
Cell Wall ◽  
Botrytis Cinerea ◽  
Defense Mechanisms ◽  
Plant Cell Wall ◽  
Quantitative Polymerase Chain Reaction ◽  
Gray Mold ◽  
Ros Generation ◽  
Genome Wide ◽  
Mold Fungus ◽  
Berry Ripening

Mature grapevine berries at the harvesting stage (MB) are very susceptible to the gray mold fungus Botrytis cinerea, while veraison berries (VB) are not. We conducted simultaneous microscopic and transcriptomic analyses of the pathogen and the host to investigate the infection process developed by B. cinerea on MB versus VB, and the plant defense mechanisms deployed to stop the fungus spreading. On the pathogen side, our genome-wide transcriptomic data revealed that B. cinerea genes upregulated during infection of MB are enriched in functional categories related to necrotrophy, such as degradation of the plant cell wall, proteolysis, membrane transport, reactive oxygen species (ROS) generation, and detoxification. Quantitative-polymerase chain reaction on a set of representative genes related to virulence and microscopic observations further demonstrated that the infection is also initiated on VB but is stopped at the penetration stage. On the plant side, genome-wide transcriptomic analysis and metabolic data revealed a defense pathway switch during berry ripening. In response to B. cinerea inoculation, VB activated a burst of ROS, the salicylate-dependent defense pathway, the synthesis of the resveratrol phytoalexin, and cell-wall strengthening. On the contrary, in infected MB, the jasmonate-dependent pathway was activated, which did not stop the fungal necrotrophic process.

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