A novel cascade allows Metarhizium robertsii to distinguish cuticle and hemocoel microenvironments during infection of insects

Pathogenic fungi precisely respond to dynamic microenvironments during infection, but the underlying mechanisms are not well understood. The insect pathogenic fungus Metarhizium robertsii is a representative fungus in which to study broad themes of fungal pathogenicity as it resembles some major plant and mammalian pathogenic fungi in its pathogenesis. Here we report on a novel cascade that regulates response of M. robertsii to 2 distinct microenvironments during its pathogenesis. On the insect cuticle, the transcription factor COH2 activates expression of cuticle penetration genes. In the hemocoel, the protein COH1 is expressed due to the reduction in epigenetic repression conferred by the histone deacetylase HDAC1 and the histone 3 acetyltransferase HAT1. COH1 interacts with COH2 to reduce COH2 stability, and this down-regulates cuticle penetration genes and up-regulates genes for hemocoel colonization. Our work significantly advances the insights into fungal pathogenicity in insects.

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Ubiquity of Insect-Derived Nitrogen Transfer to Plants by Endophytic Insect-Pathogenic Fungi: an Additional Branch of the Soil Nitrogen Cycle

Applied and Environmental Microbiology ◽

10.1128/aem.03338-13 ◽

2013 ◽

Vol 80 (5) ◽

pp. 1553-1560 ◽

Cited By ~ 88

Author(s):

Scott W. Behie ◽

Michael J. Bidochka

Keyword(s):

Panicum Virgatum ◽

Pathogenic Fungus ◽

Insect Herbivory ◽

Pathogenic Fungi ◽

Nitrogen Transfer ◽

Metarhizium Robertsii ◽

Content Type ◽

Lecanicillium Lecanii ◽

Worldwide Distribution ◽

Insect Pathogenic Fungi

ABSTRACTThe study of symbiotic nitrogen transfer in soil has largely focused on nitrogen-fixing bacteria. Vascular plants can lose a substantial amount of their nitrogen through insect herbivory. Previously, we showed that plants were able to reacquire nitrogen from insects through a partnership with the endophytic, insect-pathogenic fungusMetarhizium robertsii. That is, the endophytic capability and insect pathogenicity ofM. robertsiiare coupled so that the fungus acts as a conduit to provide insect-derived nitrogen to plant hosts. Here, we assess the ubiquity of this nitrogen transfer in fiveMetarhiziumspecies representing those with broad (M. robertsii,M. brunneum, andM. guizhouense) and narrower insect host ranges (M. acridumandM. flavoviride), as well as the insect-pathogenic fungiBeauveria bassianaandLecanicillium lecanii. Insects were injected with15N-labeled nitrogen, and we tracked the incorporation of15N into two dicots, haricot bean (Phaseolus vulgaris) and soybean (Glycine max), and two monocots, switchgrass (Panicum virgatum) and wheat (Triticum aestivum), in the presence of these fungi in soil microcosms. AllMetarhiziumspecies andB. bassianabut notL. lecaniishowed the capacity to transfer nitrogen to plants, although to various degrees. Endophytic association by these fungi increased overall plant productivity. We also showed that in the field, where microbial competition is potentially high,M. robertsiiwas able to transfer insect-derived nitrogen to plants.Metarhiziumspp. andB. bassianahave a worldwide distribution with high soil abundance and may play an important role in the ecological cycling of insect nitrogen back to plant communities.

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MrSkn7 Controls Sporulation, Cell Wall Integrity, Autolysis, and Virulence in Metarhizium robertsii

Eukaryotic Cell ◽

10.1128/ec.00266-14 ◽

2015 ◽

Vol 14 (4) ◽

pp. 396-405 ◽

Cited By ~ 17

Author(s):

Yanfang Shang ◽

Peilin Chen ◽

Yixiong Chen ◽

Yuzhen Lu ◽

Chengshu Wang

Keyword(s):

Cell Wall ◽

Target Genes ◽

Response Regulator ◽

Pathogenic Fungus ◽

Functional Divergence ◽

Pathogenic Fungi ◽

Wild Type ◽

Metarhizium Robertsii ◽

Content Type ◽

Cell Autolysis

ABSTRACT Two-component signaling pathways generally include sensor histidine kinases and response regulators. We identified an ortholog of the response regulator protein Skn7 in the insect-pathogenic fungus Metarhizium robertsii , which we named MrSkn7. Gene deletion assays and functional characterizations indicated that MrSkn7 functions as a transcription factor. The MrSkn7 null mutant of M. robertsii lost the ability to sporulate and had defects in cell wall biosynthesis but was not sensitive to oxidative and osmotic stresses compared to the wild type. However, the mutant was able to produce spores under salt stress. Insect bioassays using these spores showed that the virulence of the mutant was significantly impaired compared to that of the wild type due to the failures to form the infection structure appressorium and evade host immunity. In particular, deletion of MrSkn7 triggered cell autolysis with typical features such as cell vacuolization, downregulation of repressor genes, and upregulation of autolysis-related genes such as extracellular chitinases and proteases. Promoter binding assays confirmed that MrSkn7 could directly or indirectly control different putative target genes. Taken together, the results of this study help us understand the functional divergence of Skn7 orthologs as well as the mechanisms underlying the development and control of virulence in insect-pathogenic fungi.

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The Intermediates in Branched-Chain Amino Acid Biosynthesis Are Indispensable for Conidial Germination of the Insect-Pathogenic Fungus Metarhizium robertsii

Applied and Environmental Microbiology ◽

10.1128/aem.01682-20 ◽

2020 ◽

Vol 86 (20) ◽

Author(s):

Feifei Luo ◽

Hongxia Zhou ◽

Xue Zhou ◽

Xiangyun Xie ◽

You Li ◽

...

Keyword(s):

Amino Acid ◽

Pathogenic Fungus ◽

Pathogenic Fungi ◽

Conidial Germination ◽

Keto Acid ◽

Metarhizium Robertsii ◽

Content Type ◽

Branched Chain Amino Acid ◽

Key Enzyme ◽

Branched Chain

ABSTRACT Metarhizium spp. are well-known biocontrol agents used worldwide to control different insect pests. Keto-acid reductoisomerase (ILVC) is a key enzyme for branched-chain amino acid (BCAA) biosynthesis, and it regulates many physiological activities. However, its functions in insect-pathogenic fungi are poorly understood. In this work, we identified MrilvC in M. robertsii and dissected its roles in fungal growth, conidiation, germination, destruxin biosynthesis, environmental stress response, and insecticidal virulence. BCAA metabolism affects conidial yields and germination. However, BCAAs cannot recover the conidial germination of an MrilvC-deficient strain. Further feeding assays with intermediates showed that some conidia of the ΔMrilvC mutant start to germinate. Therefore, it is the germination defect that causes the complete failures of conidial penetration and pathogenicity in the ΔMrilvC mutant. In conclusion, we found intermediates in BCAA biosynthesis are indispensable for Metarhizium robertsii conidial germination. This study will advance our understanding of the fungal germination mechanism. IMPORTANCE Branched-chain amino acid (BCAA) metabolism plays a significant role in many biological activities beyond protein synthesis. Spore germination initiates the first stage of vegetative growth, which is critical for the virulence of pathogenic fungi. In this study, we demonstrated that the keto-acid reductoisomerase MrILVC, a key enzyme for BCAA biosynthesis, from the insect-pathogenic fungus Metarhizium robertsii is associated with conidial germination and fungal pathogenicity. Surprisingly, the germination of the ΔMrilvC mutant was restored when supplemented with the intermediates of BCAA metabolism rather than three BCAAs. The result was significantly different from that of plant-pathogenic fungi. Therefore, this report highlights that the intermediates in BCAA biosynthesis are indispensable for conidial germination of M. robertsii.

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Horizontal gene transfer allowed the emergence of broad host range entomopathogens

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1816430116 ◽

2019 ◽

Vol 116 (16) ◽

pp. 7982-7989 ◽

Cited By ~ 18

Author(s):

Qiangqiang Zhang ◽

Xiaoxuan Chen ◽

Chuan Xu ◽

Hong Zhao ◽

Xing Zhang ◽

...

Keyword(s):

Gene Transfer ◽

Horizontal Gene Transfer ◽

Host Range ◽

Range Expansion ◽

Pathogenic Fungi ◽

Broad Host Range ◽

Metarhizium Robertsii ◽

Metarhizium Acridum ◽

Host Range Expansion ◽

Underlying Mechanisms

The emergence of new pathogenic fungi has profoundly impacted global biota, but the underlying mechanisms behind host shifts remain largely unknown. The endophytic insect pathogen Metarhizium robertsii evolved from fungi that were plant associates, and entomopathogenicity is a more recently acquired adaptation. Here we report that the broad host-range entomopathogen M. robertsii has 18 genes that are derived via horizontal gene transfer (HGT). The necessity of degrading insect cuticle served as a major selective pressure to retain these genes, as 12 are up-regulated during penetration; 6 were confirmed to have a role in penetration, and their collective actions are indispensable for infection. Two lipid-carrier genes are involved in utilizing epicuticular lipids, and a third (MrNPC2a) facilitates hemocoel colonization. Three proteases degraded the procuticular protein matrix, which facilitated up-regulation of other cuticle-degrading enzymes. The three lipid carriers and one of the proteases are present in all analyzed Metarhizium species and are essential for entomopathogenicity. Acquisition of another protease (MAA_01413) in an ancestor of broad host-range lineages contributed to their host-range expansion, as heterologous expression in the locust specialist Metarhizium acridum enabled it to kill caterpillars. Our work reveals that HGT was a key mechanism in the emergence of entomopathogenicity in Metarhizium from a plant-associated ancestor and in subsequent host-range expansion by some Metarhizium lineages.

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New Spirocyclic Hydroxamic Acids as Effective Antiproliferative Agents

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520620666200527132420 ◽

2020 ◽

Vol 20 ◽

Cited By ~ 1

Author(s):

Margarita E. Neganova ◽

Sergey G. Klochkov ◽

Yulia R. Aleksandrova ◽

Vladimir N. Osipov ◽

Dmitry V. Avdeev ◽

...

Keyword(s):

Tumor Cell ◽

Histone Deacetylase ◽

Cell Lines ◽

Anticancer Agents ◽

Antioxidant Activities ◽

Hydroxamic Acids ◽

Cytotoxic Activities ◽

Tumor Cell Lines ◽

Hek 293 ◽

Underlying Mechanisms

Aims: The main goal of this work where is to synthesize new original spirocyclic hydroxamic acids, investigate their cytotoxicity against to the panel of tumor cell lines and possible mechanism of action of these active compounds. Background: Hydroxamic acids are one of the promising classes of chemical compounds with proven has anticancer potential properties. This is manifested in the presence of metal chelating and antioxidant activities, the ability to inhibit histone deacetylase enzymes and a chemosensitizing effect against well known cytostatics. Objective: Original spirocyclic hydroxamic acids were synthesized and spectrums of their antiproliferative activities were investigated. Methods: The cytotoxic activities on different tumor lines (SH-SY5Y, HeLa and healthy cells HEK-293) were investigated and determined possible underlying mechanisms of their activity. Result: New original spirocyclic hydroxamic acids were synthesized. These compounds exhibit antiproliferative properties against of the various tumor cultures cells and also exhibits antioxidant activity, a depolarizing effect on the mitochondrial membrane, inhibit the activity of the histone deacetylase enzyme, and also decrease of basal glycolysis and glycolytic capacity reserve of HeLa and SH-SY5Y tumor cell lines. Conclusion: The most promising are compounds 5j-l containing two chlorine atoms as substituents in the quinazoline part of the molecule and hydroxamate function. Therefore, these compounds can be considered as hit compounds for the development on their basis multi-target anticancer agents.

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A Gene Related to Yeast HOS2 Histone Deacetylase Affects Extracellular Depolymerase Expression and Virulence in a Plant Pathogenic Fungus

The Plant Cell ◽

10.1105/tpc.13.7.1609 ◽

2001 ◽

Vol 13 (7) ◽

pp. 1609-1624 ◽

Cited By ~ 26

Author(s):

D. Baidyaroy

Keyword(s):

Histone Deacetylase ◽

Pathogenic Fungus ◽

Plant Pathogenic Fungus

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Brassinosteroids repress the seed maturation program during the seed-to-seedling transition

Plant Physiology ◽

10.1093/plphys/kiab089 ◽

2021 ◽

Author(s):

Jiuxiao Ruan ◽

Huhui Chen ◽

Tao Zhu ◽

Yaoguang Yu ◽

Yawen Lei ◽

...

Keyword(s):

Arabidopsis Thaliana ◽

Transcription Factor ◽

Abscisic Acid ◽

Plant Hormone ◽

Flowering Plants ◽

Seed Maturation ◽

Domain Protein ◽

Abscisic Acid Insensitive3 ◽

Underlying Mechanisms ◽

Embryonic Structure

Abstract In flowering plants, repression of the seed maturation program is essential for the transition from the seed to the vegetative phase, but the underlying mechanisms remain poorly understood. The B3-domain protein VIVIPAROUS1/ABSCISIC ACID-INSENSITIVE3-LIKE 1 (VAL1) is involved in repressing the seed maturation program. Here we uncovered a molecular network triggered by the plant hormone brassinosteroid (BR) that inhibits the seed maturation program during the seed-to-seedling transition in Arabidopsis (Arabidopsis thaliana). val1-2 mutant seedlings treated with a BR biosynthesis inhibitor form embryonic structures, whereas BR signaling gain-of-function mutations rescue the embryonic structure trait. Furthermore, the BR-activated transcription factors BRI1-EMS-SUPPRESSOR 1 and BRASSINAZOLE-RESISTANT 1 bind directly to the promoter of AGAMOUS-LIKE15 (AGL15), which encodes a transcription factor involved in activating the seed maturation program, and suppress its expression. Genetic analysis indicated that BR signaling is epistatic to AGL15 and represses the seed maturation program by downregulating AGL15. Finally, we showed that the BR-mediated pathway functions synergistically with the VAL1/2-mediated pathway to ensure the full repression of the seed maturation program. Together, our work uncovered a mechanism underlying the suppression of the seed maturation program, shedding light on how BR promotes seedling growth.

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Metabolome and proteome analyses reveal transcriptional misregulation in glycolysis of engineered E. coli

Nature Communications ◽

10.1038/s41467-021-25142-0 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Chun-Ying Wang ◽

Martin Lempp ◽

Niklas Farke ◽

Stefano Donati ◽

Timo Glatter ◽

...

Keyword(s):

Transcription Factor ◽

Binding Site ◽

Metabolic Pathways ◽

Glycerol Production ◽

Inducible Promoters ◽

E Coli ◽

Inhibition Of Glycolysis ◽

Underlying Mechanisms ◽

Rate Limiting ◽

Engineered Bacteria

AbstractSynthetic metabolic pathways are a burden for engineered bacteria, but the underlying mechanisms often remain elusive. Here we show that the misregulated activity of the transcription factor Cra is responsible for the growth burden of glycerol overproducing E. coli. Glycerol production decreases the concentration of fructose-1,6-bisphoshate (FBP), which then activates Cra resulting in the downregulation of glycolytic enzymes and upregulation of gluconeogenesis enzymes. Because cells grow on glucose, the improper activation of gluconeogenesis and the concomitant inhibition of glycolysis likely impairs growth at higher induction of the glycerol pathway. We solve this misregulation by engineering a Cra-binding site in the promoter controlling the expression of the rate limiting enzyme of the glycerol pathway to maintain FBP levels sufficiently high. We show the broad applicability of this approach by engineering Cra-dependent regulation into a set of constitutive and inducible promoters, and use one of them to overproduce carotenoids in E. coli.

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Glycoconjugates on the surface of spores of the pathogenic fungus Phytophthora cinnamomi studied using fluorescence and electron microscopy and flow cytometry

Canadian Journal of Microbiology ◽

10.1139/m90-032 ◽

1990 ◽

Vol 36 (3) ◽

pp. 183-192 ◽

Cited By ~ 22

Author(s):

A. R. Hardham ◽

E. Suzaki

Keyword(s):

Flow Cytometry ◽

Plasma Membrane ◽

Cell Surface ◽

Phytophthora Cinnamomi ◽

Pathogenic Fungus ◽

Fluorescein Isothiocyanate ◽

Pathogenic Fungi ◽

Surface Flow ◽

Soybean Agglutinin ◽

Binding Material

Glycoconjugates on the surface of zoospores and cysts of the pathogenic fungus Phytophthora cinnamomi have been studied using fluorescein isothiocyanate labelled lectins for fluorescence microscopy and flow cytometry, and ferritin- and gold-labelled lectins for ultrastructural analysis. Of the five lectins used, only concanavalin A (ConA) binds to the surface of the zoospores, including the flagella and water expulsion vacuole. This suggests that of accessible saccharides, glucosyl or mannosyl residues predominate on the outer surface of the zoospore plasma membrane. Early in encystment, a system of flat disc-like cisternae, which underlie the zoospore plasma membrane, vesiculate. These and other small peripheral vesicles quickly disappear. After the induction of encystment, ConA is no longer localised close to the plasma membrane but binds to material loosely associated with the cell surface. Quantitative measurements by flow cytometry indicate that the ConA-binding material is gradually lost from the cell surface. The cyst wall is weakly labelled, but the site of germ tube emergence stains intensely. During the first 2 min after the induction of encystment, material that binds soybean agglutinin, Helix pommatia agglutinin, and peanut agglutinin appears on the surface of the fungal cells. The distribution of this material, rich in galactosyl or N-acetyl-D-galactosaminosyl residues, is initially patchy, but by 5 min the material evenly coats most of the cell surface. Labelling of zoospores in which intracellular sites are accessible indicates that the soybean agglutinin binding material is stored in vesicles that lie beneath the plasma membrane. Quantitation of soybean agglutinin labelling shows that maximum binding occurs 2–3 min after the induction of encystment. Key words: cell surface, flow cytometry, lectins, pathogenic fungi, Phytophthora cinnamomi.

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