ABSTRACT
The lymphatic circulation plays a crucial role in maintaining fluid balance and supporting immune responses by returning serum proteins and lipids to the systemic circulation. Lymphatic leaks, though rare, pose significant challenges post-radical neck surgery, oesophagectomy, and thoracic or retroperitoneal oncological resections, leading to heightened morbidity and mortality. Managing lymphatic leaks necessitates consideration of aetiology, severity, and volume of leakage. Traditionally, treatment involved conservative measures such as dietary restrictions, drainage, and medical management, with surgical intervention reserved for severe cases, albeit with variable outcomes and extended recovery periods. Lymphography, introduced in the 1950s, initially served as a diagnostic tool for lymphoedema, lymphoma, tumour staging, and monitoring chemotherapy response. However, its widespread adoption was impeded by alternative techniques like Computed Tomography, learning curves, and its associated complications. Contemporary lymphatic interventions have evolved, favouring nodal lymphangiography over pedal lymphangiography for its technical simplicity and reduced complexity. Effective management of chylous leaks mandates a multimodal approach encompassing clinical evaluation and imaging techniques. In cases where conservative management proves ineffective, embolization through conventional lymphangiography by bipedal dissection or intranodal injection emerges as a viable option. This review underscores the importance of a comprehensive approach to diagnosing and treating lymphatic leaks, highlighting advancements in imaging and therapeutic interventions that enhance patient outcomes.
ABSTRACT
Trichoderma virens is a beneficial fungus that helps plants fight pathogens and abiotic stresses and thereby enhances crop yields. Unlike other Trichoderma spp., there are two well-defined strains (P and Q) of T. virens, classified by secondary metabolites profiling, primarily the biosynthesis of the nonribosomal, strong antimicrobial agents gliotoxin (Q) and gliovirin (P). We have studied the phenotypic and biocontrol properties of two well-studied representative isolates (T. virens Gv29-8 and T. virens GvW/IMI304061) that represent a Q strain and a P strain of T. virens, respectively. We refined the genome assembly of the P strain using nanopore technology, and we compared it with the Q strain. The differences between the genomes include gene expansion in the Q strain. T. virens Gv29-8 is weaker than GvW as a mycoparasite on the broad host-range plant pathogen Sclerotium rolfsii, and it is ineffective as a biocontrol agent when applied to pathogen-infested soil. T. virens Gv29-8 proved to be phytotoxic to Arabidopsis seedlings, whereas the effect of T. virens GvW was not major. Both strains colonized the surface and outer cortex layer of tomato roots, with about 40% higher colonization by T. virens Gv29-8. T. virens Gv29-8 induced the expression of a larger set of tomato genes than did T. virens GvW, although some tomato genes were uniquely induced in response to T. virens GvW. We studied the comparative transcriptome response of T. virens Gv29-8 and T. virens GvW to S. rolfsii. A larger set of genes was regulated in T. virens GvW than in T. virens Gv29-8 in the presence of the plant pathogen. IMPORTANCE Trichoderma virens populations that were earlier classified into two strains (P and Q) based on secondary metabolites profiling are also phenotypically and genetically distinct, with the latter being ineffective in controlling the devastating, broad host range plant pathogen Sclerotium rolfsii. The two strains also provoke distinct as well as overlapping transcriptional responses to the presence of the plant and the pathogen. This study enriches our knowledge of Trichoderma-plant-pathogen interactions and identifies novel candidate genes for further research and deployment in agriculture.
ABSTRACT
Beneficial interaction of members of the fungal genus Trichoderma with plant roots primes the plant immune system, promoting systemic resistance to pathogen infection. Some strains of Trichoderma virens produce gliotoxin, a fungal epidithiodioxopiperazine (ETP)-type secondary metabolite that is toxic to animal cells. It induces apoptosis, prevents NF-κB activation via the inhibition of the proteasome, and has immunosuppressive properties. Gliotoxin is known to be involved in the antagonism of rhizosphere microorganisms. To investigate whether this metabolite has a role in the interaction of Trichoderma with plant roots, we compared gliotoxin-producing and nonproducing T. virens strains. Both colonize the root surface and outer layers, but they have differential effects on root growth and architecture. The responses of tomato plants to a pathogen challenge were followed at several levels: lesion development, levels of ethylene, and reactive oxygen species. The transcriptomic signature of the shoot tissue in response to root interaction with producing and nonproducing T. virens strains was monitored. Gliotoxin producers provided stronger protection against foliar pathogens, compared to nonproducing strains. This was reflected in the transcriptomic signature, which showed the induction of defense-related genes. Two markers of plant defense response, PR1 and Pti-5, were differentially induced in response to pure gliotoxin. Gliotoxin thus acts as a microbial signal, which the plant immune system recognizes, directly or indirectly, to promote a defense response. IMPORTANCE A single fungal metabolite induces far-reaching transcriptomic reprogramming in the plant, priming immune responses and defense, in contrast to its immunosuppressive effect on animal cells. While the negative effects of gliotoxin-producing Trichoderma strains on growth may be observed only under a particular set of laboratory conditions, gliotoxin-linked molecular patterns, including the potential for limited cell death, could strongly prime plant defense, even in mature soil-grown plants in which the same Trichoderma strain promotes growth.
Subject(s)
Gliotoxin , Hypocrea , Solanum lycopersicum , Trichoderma , Animals , Hypocrea/metabolism , Solanum lycopersicum/microbiology , Plant Diseases/microbiology , Plant Immunity , Plant Roots/microbiology , Trichoderma/genetics , Trichoderma/metabolismABSTRACT
Fungal spores, germlings, and mycelia adhere to substrates, including host tissues. The adhesive forces depend on the substrate and on the adhesins, the fungal cell surface proteins. Attachment is often a prerequisite for the invasion of the host, hence its importance. Adhesion visibly precedes colonization of root surfaces and outer cortex layers, but little is known about the molecular details. We propose that by starting from what is already known from other fungi, including yeast and other filamentous pathogens and symbionts, the mechanism and function of Trichoderma adhesion will become accessible. There is a sequence, and perhaps functional, homology to other rhizosphere-competent Sordariomycetes. Specifically, Verticillium dahliae is a soil-borne pathogen that establishes itself in the xylem and causes destructive wilt disease. Metarhizium species are best-known as insect pathogens with biocontrol potential, but they also colonize roots. Verticillium orthologs of the yeast Flo8 transcription factor, Som1, and several other relevant genes are already under study for their roles in adhesion. Metarhizium encodes relevant adhesins. Trichoderma virens encodes homologs of Som1, as well as adhesin candidates. These genes should provide exciting leads toward the first step in the establishment of beneficial interactions with roots in the rhizosphere.
ABSTRACT
Development of regulated cellular processes and signaling methods in synthetic cells is essential for their integration with living materials. Light is an attractive tool to achieve this, but the limited penetration depth into tissue of visible light restricts its usability for in-vivo applications. Here, we describe the design and implementation of bioluminescent intercellular and intracellular signaling mechanisms in synthetic cells, dismissing the need for an external light source. First, we engineer light generating SCs with an optimized lipid membrane and internal composition, to maximize luciferase expression levels and enable high-intensity emission. Next, we show these cells' capacity to trigger bioprocesses in natural cells by initiating asexual sporulation of dark-grown mycelial cells of the fungus Trichoderma atroviride. Finally, we demonstrate regulated transcription and membrane recruitment in synthetic cells using bioluminescent intracellular signaling with self-activating fusion proteins. These functionalities pave the way for deploying synthetic cells as embeddable microscale light sources that are capable of controlling engineered processes inside tissues.
Subject(s)
Artificial Cells , Optogenetics , Light , Luciferases , Optogenetics/methods , Signal TransductionABSTRACT
BACKGROUND: G protein-coupled receptors (GPCRs) usually regulate cellular processes via activation of intracellular signaling pathways. However, we have previously shown that in several cell lines, GqPCRs induce immediate inactivation of the AKT pathway, which leads to JNK-dependent apoptosis. This apoptosis-inducing AKT inactivation is essential for physiological functions of several GqPCRs, including those for PGF2α and GnRH. METHODS: Here we used kinase activity assays of PI3K and followed phosphorylation state of proteins using specific antibodies. In addition, we used coimmunoprecipitation and proximity ligation assays to follow protein-protein interactions. Apoptosis was detected by TUNEL assay and PARP1 cleavage. RESULTS: We identified the mechanism that allows the unique stimulated inactivation of AKT and show that the main regulator of this process is the phosphatase PP2A, operating with the non-canonical regulatory subunit IGBP1. In resting cells, an IGBP1-PP2Ac dimer binds to PI3K, dephosphorylates the inhibitory pSer608-p85 of PI3K and thus maintains its high basal activity. Upon GqPCR activation, the PP2Ac-IGBP1 dimer detaches from PI3K and thus allows the inhibitory dephosphorylation. At this stage, the free PP2Ac together with IGBP1 and PP2Aa binds to AKT, causing its dephosphorylation and inactivation. CONCLUSION: Our results show a stimulated shift of PP2Ac from PI3K to AKT termed "PP2A switch" that represses the PI3K/AKT pathway, providing a unique mechanism of GPCR-stimulated dephosphorylation. Video Abstract.
Subject(s)
Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , Phosphatidylinositol 3-Kinases/metabolism , Phosphorylation , Proto-Oncogene Proteins c-akt/metabolism , Receptors, G-Protein-Coupled/metabolism , Signal TransductionABSTRACT
The establishment of plant-fungus mutualistic interaction requires bidirectional molecular crosstalk. Therefore, the analysis of the interacting organisms secretomes would help to understand how such relationships are established. Here, a gel-free shotgun proteomics approach was used to identify the secreted proteins of the plant Arabidopsis thaliana and the mutualistic fungus Trichoderma atroviride during their interaction. A total of 126 proteins of Arabidopsis and 1027 of T. atroviride were identified. Among them, 118 and 780 were differentially modulated, respectively. Bioinformatic analysis unveiled that both organisms' secretomes were enriched with enzymes. In T. atroviride, glycosidases, aspartic endopeptidases, and dehydrogenases increased in response to Arabidopsis. Additionally, amidases, protein-serine/threonine kinases, and hydro-lyases showed decreased levels. Furthermore, peroxidases, cysteine endopeptidases, and enzymes related to the catabolism of secondary metabolites increased in the plant secretome. In contrast, pathogenesis-related proteins and protease inhibitors decreased in response to the fungus. Notably, the glutamate:glyoxylate aminotransferase GGAT1 was secreted by Arabidopsis during its interaction with T. atroviride. Our study showed that GGAT1 is partially required for plant growth stimulation and on the induction of the plant systemic resistance by T. atroviride. Additionally, GGAT1 seems to participate in the negative regulation of the plant systemic resistance against B. cinerea through a mechanism involving H2O2 production.
Subject(s)
Arabidopsis/metabolism , Arabidopsis/microbiology , Botrytis , Disease Resistance , Host-Pathogen Interactions , Metabolomics , Plant Diseases/microbiology , Trichoderma , Computational Biology/methods , Glutamic Acid/metabolism , Metabolomics/methods , Phenotype , Plant Development , Symbiosis , Transaminases/genetics , Transaminases/metabolismABSTRACT
Protein phosphorylation cascades are universal in cell signaling. While kinome diversity allows specific phosphorylation events, relatively few phosphatases dephosphorylate key signaling proteins. Fungal mitogen activated protein kinases (MAPK), in contrast to their mammalian counterparts, often show detectable basal phosphorylation levels. Dephosphorylation, therefore, could act as a signal. In Cochliobolus heterostrophus, the Dothideomycete causing Southern corn leaf blight, ferulic acid (FA)-an abundant phenolic found in plant host cell walls-acts as a signal to rapidly dephosphorylate the stress-activated MAP kinase Hog1 (High Osmolarity Glycerol 1). In order to identify the protein phosphatases responsible, we constructed mutants in Hog1 phosphatases predicted from the genome by homology to yeast and other species. We found that Cochliobolus heterostrophus mutants lacking PtcB, a member of the PP2C family, exhibited altered growth, sporulation, and attenuated dephosphorylation in response to FA. The loss of the dual-specificity phosphatase CDC14 led to slow growth, decreased virulence, and attenuated dephosphorylation. Mutants in two predicted tyrosine phosphatase genes PTP1 and PTP2 showed normal development and virulence. Our results suggest that a network of phosphatases modulate Hog1's dual phosphorylation levels. The mutants we constructed in this work provide a starting point to further unravel the signaling hierarchy by which exposure to FA leads to stress responses in the pathogen.
ABSTRACT
Trichoderma virens is a well-known mycoparasitic fungal symbiont that is valued for its biocontrol capabilities. T. virens initiates a symbiotic relationship with a plant host through the colonization of its roots. To achieve colonization, the fungus must communicate with the host and evade its innate defenses. In this study, we explored the genes involved with the host communication and colonization process through transcriptomic profiling of the wild-type fungus and selected deletion mutants as they colonized maize roots. Transcriptome profiles of the T. virens colonization of maize roots over time revealed that 24 h post inoculation appeared to be a key time for plant-microbe communication, with many key gene categories, including signal transduction mechanisms and carbohydrate transport and metabolism, peaking in expression at this early colonization time point. The transcriptomic profiles of Sm1 and Sir1 deletion mutants in the presence of plants demonstrated that Sir1, rather than Sm1, appears to be the key regulator of the fungal response to maize, with 64% more unique differentially expressed genes compared to Sm1. Additionally, we developed a novel algorithm utilizing gene clustering and coexpression network analyses to select potential colonization-related gene targets for characterization. About 40% of the genes identified by the algorithm would have been missed using previous methods for selecting gene targets.
ABSTRACT
The anticancer antibiotic heptelidic acid is a sesquiterpene lactone produced by the beneficial plant fungus Trichoderma virens. This species has been separated into two strains, referred to as P and Q, based on its biosynthesis of secondary metabolites; notably, only P-strains were reported to produce heptelidic acid. While characterizing a Q-strain of T. virens containing a directed mutation in the non-ribosomal peptide synthetase encoding gene Tex7, the appearance of an unknown compound in anomalously large quantities was visualized by TLC. Using a combination of HPLC, LC-MS/MS, and NMR spectroscopy, this compound was identified as heptelidic acid. This discovery alters the strain classification structure of T. virens. Additionally, the Tex7 mutants inhibited growth of maize seedlings, while retaining the ability to induce systemic resistance against the foliar fungal pathogen, Cochliobolus heterostrophus.
Subject(s)
Antibiotics, Antineoplastic/metabolism , Fungal Proteins/genetics , Peptide Synthases/genetics , Trichoderma/genetics , Fungal Proteins/metabolism , Gene Deletion , Genes, Fungal , Multigene Family , Peptide Synthases/metabolism , Sesquiterpenes/metabolism , Trichoderma/metabolism , Zea mays/growth & development , Zea mays/microbiologySubject(s)
Bronchitis/therapy , Embolization, Therapeutic , Magnetic Resonance Imaging , Thoracic Duct/diagnostic imaging , Ultrasonography , Adolescent , Bronchitis/diagnostic imaging , Female , Humans , Image Interpretation, Computer-Assisted , Multimodal Imaging , Predictive Value of Tests , Punctures , Treatment OutcomeABSTRACT
Fungal pathogens need to contend with stresses including oxidants and antimicrobial chemicals resulting from host defenses. ChAP1 of Cochliobolus heterostrophus, agent of Southern corn leaf blight, encodes an ortholog of yeast YAP1. ChAP1 is retained in the nucleus in response to plant-derived phenolic acids, in addition to its well-studied activation by oxidants. Here, we used transcriptome profiling to ask which genes are regulated in response to ChAP1 activation by ferulic acid (FA), a phenolic abundant in the maize host. Nuclearization of ChAP1 in response to phenolics is not followed by strong expression of genes needed for oxidative stress tolerance. We, therefore, compared the transcriptomes of the wild-type pathogen and a ChAP1 deletion mutant, to study the function of ChAP1 in response to FA. We hypothesized that if ChAP1 is retained in the nucleus under plant-related stress conditions yet in the absence of obvious oxidant stress, it should have additional regulatory functions. The transcriptional signature in response to FA in the wild type compared to the mutant sheds light on the signaling mechanisms and response pathways by which ChAP1 can mediate tolerance to ferulic acid, distinct from its previously known role in the antioxidant response. The ChAP1-dependent FA regulon consists mainly of two large clusters. The enrichment of transport and metabolism-related genes in cluster 1 indicates that C. heterostrophus degrades FA and removes it from the cell. When this fails at increasing stress levels, FA provides a signal for cell death, indicated by the enrichment of cell death-related genes in cluster 2. By quantitation of survival and by TUNEL assays, we show that ChAP1 promotes survival and mitigates cell death. Growth rate data show a time window in which the mutant colony expands faster than the wild type. The results delineate a transcriptional regulatory pattern in which ChAP1 helps balance a survival response for tolerance to FA, against a pathway promoting cell death in the pathogen. A general model for the transition from a phase where the return to homeostasis dominates to a phase leading to the onset of cell death provides a context for understanding these findings.
Subject(s)
Ascomycota/physiology , Fungal Proteins/metabolism , Host-Pathogen Interactions , Phenols/metabolism , Plant Diseases/microbiology , Transcription Factor AP-1/metabolism , Zea mays/microbiology , Biomarkers , Computational Biology/methods , Gene Expression Profiling , Gene Expression Regulation , Gene OntologyABSTRACT
Resumen El hallazgo de hemoperitoneo en el post parto secundario a la rotura aneurismática de la arteria ovárica es una situación clínica extremadamente rara que presenta un cuadro clínico inespecífico y puede poner en riesgo la vida del paciente. El ultrasonido es una modalidad segura y rápida para la detección de líquido libre intraperitoneal. (1) . La tomografía computada es la herramienta de elección para un diagnóstico rápido y seguro (2) ; y la angiografía con embolización durante el mismo procedimiento es una alternativa útil y altamente efectiva para la resolución del cuadro. (3). Presentamos el caso de una multípara puérpera de 34 años que consulta en el servicio de urgencia por intenso dolor abdominal. La paciente se encontraba hemodinámicamente estable y afebril. La tomografía computada demostró un hematoma retroperitoneal y hemoperitoneo asociado a un aneurisma de la arteria ovárica derecha. Fue evaluada por el servicio de radiología intervencional y se trasladó de emergencia al pabellón angiográfico donde se realizó la embolización de la lesión mediante la cateterización vascular supra selectiva. La paciente evolucionó de manera favorable y fue dada de alta una semana después. Es necesario tener un alto índice de sospecha en pacientes de riesgo para lograr un diagnóstico y tratamiento oportuno.
SUMMARY Spontaneous ovarian artery aneurysm rupture is a rare postpartum life-threatening event with non-specific clinical manifestations. The present article reports the case of a 34 year old multiparous post partum women who came to the emergency department with acute onset of intense abdominal right flank pain. Patient was afebrile and hemodynamically stable. A computed tomography revealed a retroperitoneal haematoma and hemoperitoneum related to an aneurysm of the right ovarian artery. The patient was taken to the interventional radiology suite and selective embolization was performed. Following the procedure, the patient symptoms subsided and 7 days later she was discharged. A high index of suspicion in patients with risk factors can lead to a prompt diagnosis and treatment. Computed tomography is the image modality for a fast and safe evaluation, although diagnostic angiography and subsequent transcatheter embolization are thought to be effective for treatment.
Subject(s)
Humans , Female , Adult , Aneurysm, Ruptured/complications , Aneurysm, Ruptured/diagnostic imaging , Postpartum Period , Hemoperitoneum , Ovary/blood supply , Rupture, Spontaneous , Tomography, X-Ray Computed , Uterine Artery EmbolizationABSTRACT
Host defenses expose fungal pathogens to oxidants and antimicrobial chemicals. The fungal cell employs conserved eukaryotic signaling pathways and dedicated transcription factors to program its response to these stresses. The oxidant-sensitive transcription factor of yeast, YAP1, and its orthologs in filamentous fungi, are central to tolerance to oxidative stress. The C-terminal domain of YAP1 contains cysteine residues that, under oxidizing conditions, form an intramolecular disulfide bridge locking the molecule in a conformation where the nuclear export sequence is masked. YAP1 accumulates in the nucleus, promoting transcription of genes that provide the cell with the ability to counteract oxidative stress. Chemicals including xenobiotics and plant signals can also promote YAP1 nuclearization in yeast and filamentous fungi. This could happen via direct or indirect oxidative stress, or by a different biochemical pathway. Plant phenolics are known antioxidants, yet they have been shown to elicit cellular responses that would usually be triggered to counter oxidant stress. Here we will discuss the evidence that YAP1 and MAPK pathways respond to phenolic compounds. Following this and other examples, we explore here how oxidative-stress sensing networks of fungi might have evolved to detect chemical stressors. Furthermore, we draw functional parallels between fungal YAP1 and mammalian Keap1-Nrf2 signaling systems.
ABSTRACT
Trichoderma spp. are widely used as commercial biofungicides, and most commercial formulations are conidia based. Identification of genes that regulate conidiation would thus be of help in genetic reprogramming of these species to optimize sporulation. In this study, we constructed an SSH (suppression subtractive hybridization) library from RNA samples of the wild type strain and a non-conidiating mutant, M7, grown under constant illumination for 2 days. We identified several genes that are underexpressed in the mutant. Some of these genes are related to secondary metabolism, and a few could be associated with conidiation. Genes coding for the following proteins, among others, were identified: O-methyl transferase, ATPase, alpha/beta-hydrolase, WD repeat containing protein, dehydrogenase, thioesterase, translationally controlled tumour protein, and a proline-glycine-tyrosine-rich protein (PGYRP) that has been annotated in T.reesei as a signalling protein. Two of these genes, encoding Pgy1, a novel PGYRP, and Ecm33, a GPI-anchored cell wall protein, were further studied in detail by generation of deletion mutants. We demonstrate here that both these genes not only regulate radial growth and conidiation in Trichoderma virens, but are also involved in antagonism against soil-borne wide host range plant pathogens. Furthermore, deletion of ecm33 affected hydrophobicity and cell wall integrity.
Subject(s)
Cell Wall/genetics , Fungal Proteins/genetics , Soil Microbiology , Trichoderma/genetics , Gene Expression Regulation, Fungal , Gene Library , Glycine/genetics , Proline/genetics , RNA, Fungal/genetics , Spores, Fungal/genetics , Spores, Fungal/growth & development , Tyrosine/geneticsABSTRACT
Plant aromatic compounds provide signals and a nutrient source to pathogens, and also act as stressors. Structure-activity relationships suggest two pathways sensing these compounds in the maize pathogen Cochliobolus heterostrophus, one triggering a stress response, and one inducing enzymes for their degradation. Focusing on the stress pathway, we found that ferulic acid causes rapid appearance of TUNEL-positive nuclei, dispersion of histone H1:GFP, hyphal shrinkage, and eventually membrane damage. These hallmarks of programmed cell death (PCD) were not seen upon exposure to caffeic acid, a very similar compound. Exposure to ferulic acid dephosphorylated two MAP kinases: Hog1 (stress activated) and Chk1 (pathogenicity related), while increasing phosphorylation of Mps1 (cell integrity related). Mutants lacking Hog1 or Chk1 are hypersensitive to ferulic acid while Mps1 mutants are not. These results implicate three MAPK pathways in the stress response. Ferulic acid and the antifungal fludioxonil have opposite additive effects on survival and on dephosphorylation of Hog1, which is thus implicated in survival. The results may explain why some fungal pathogens of plants undergo cell death early in host invasion, when phenolics are released from plant tissue.
Subject(s)
Apoptosis , Ascomycota/cytology , Fungal Proteins/metabolism , Hydroxybenzoates/metabolism , Mitogen-Activated Protein Kinases/metabolism , Plant Diseases/microbiology , Zea mays/metabolism , Ascomycota/genetics , Ascomycota/metabolism , Fungal Proteins/genetics , Gene Expression Regulation, Fungal , Host-Pathogen Interactions , Mitogen-Activated Protein Kinases/genetics , Phosphorylation , Signal Transduction , Zea mays/microbiologyABSTRACT
The biocontrol agent, Trichoderma virens, has the ability to protect plants from pathogens by eliciting plant defense responses, involvement in mycoparasitism, or secreting antagonistic secondary metabolites. SM1, an elicitor of induced systemic resistance (ISR), was found to have three paralogs within the T. virens genome. The paralog sm2 is highly expressed in the presence of plant roots. Gene deletion mutants of sm2 were generated and the mutants were found to overproduce SM1. The ability to elicit ISR in maize against Colletotrichum graminicola was not compromised for the mutants compared to that of wild type isolate. However, the deletion strains had a significantly lowered ability to colonize maize roots. This appears to be the first report on the involvement of an effector-like protein in colonization of roots by Trichoderma.
Subject(s)
Fungal Proteins/metabolism , Plant Roots/microbiology , Trichoderma/growth & development , Zea mays/microbiology , Fungal Proteins/genetics , Gene Deletion , Gene Expression Profiling , Plant Roots/immunology , Trichoderma/genetics , Zea mays/immunologyABSTRACT
Trichoderma virens is a biocontrol agent used in agriculture to antagonize pathogens of crop plants. In addition to direct mycoparasitism of soil-borne fungal pathogens, T. virens interacts with roots. This interaction induces systemic resistance (ISR), which reduces disease in above-ground parts of the plant. In the molecular dialog between fungus and plant leading to ISR, proteins secreted by T. virens provide signals. Only a few such proteins have been characterized previously. To study the secretome, proteins were characterized from hydroponic culture systems with T. virens alone or with maize seedlings, and combined with a bioassay for ISR in maize leaves infected by the pathogen Cochliobolus heterostrophus. The secreted protein fraction from coculture of maize roots and T. virens (Tv+M) was found to have a higher ISR activity than from T. virens grown alone (Tv). A total of 280 fungal proteins were identified, 66 showing significant differences in abundance between the two conditions: 32 were higher in Tv+M and 34 were higher in Tv. Among the 34 found in higher abundance in Tv and negatively regulated by roots were 13 SSCPs (small, secreted, cysteine rich proteins), known to be important in the molecular dialog between plants and fungi. The role of four SSCPs in ISR was studied by gene knockout. All four knockout lines showed better ISR activity than WT without affecting colonization of maize roots. Furthermore, the secreted protein fraction from each of the mutant lines showed improved ISR activity compared with WT. These SSCPs, apparently, act as negative effectors reducing the defense levels in the plant and may be important for the fine tuning of ISR by Trichoderma. The down-regulation of SSCPs in interaction with plant roots implies a revision of the current model for the Trichoderma-plant symbiosis and its induction of resistance to pathogens.