The Mitogen-Activated Protein Kinase Gene Crmapk Is Involved in Clonostachys chloroleuca Mycoparasitism

Abstract

Clonostachys chloroleuca is a mycoparasite used for biocontrol of numerous fungal plant pathogens. Sequencing of the transcriptome of C. chloroleuca following mycoparasitization of the sclerotia of Sclerotinia sclerotiorum revealed significant upregulation of a mitogen-activated protein kinase (MAPK)-encoding gene, crmapk. Although MAPKs are known to regulate fungal growth and development, the function of crmapk in C. chloroleuca mycoparasitism is unclear. In this study, we investigated the role of crmapk in C. chloroleuca mycoparasitism through gene knockout and complementation. Deletion of crmapk had no influence on the C. chloroleuca morphological characteristics but could significantly reduce the mycoparasitic ability to sclerotia and biocontrol capacity to soybean Sclerotinia stem rot; crmapk complementation restored these abilities. Transcriptome analysis between Δcrmapk and the wild-type strain revealed numerous genes were significantly down-regulated after crmapk deletion, including cytochrome P450, transporters, and cell wall–degrading enzymes (CWDEs). Our findings indicate that crmapk influences C. chloroleuca mycoparasitism by regulation of genes controlling the activity of CWDEs or antibiotic production. This study provides a basis for further studies of the molecular mechanism of C. chloroleuca mycoparasitism.

Screening and characterisation of proteins interacting with the mitogen-activated protein kinase Crmapk in the fungus Clonostachys chloroleuca

“…To investigate mycoparasitism-related genes in C. chloroleuca , the transcriptome of the strain 67-1 associated with S. sclerotiorum sclerotia was sequenced and the differentially expressed genes 8 h, 24 h and 48 h after sclerotia induction were analysed 9 . Combining with these results, we found that 38 interacting genes were differentially expressed (|Log 2 FC| ≥ 1 and p ≤ 0.05; Table 1 ), among which 15 genes ( NODE_15_2 , NODE_171_30 , NODE_176_9 , NODE_198_20 , NODE_2_38 , NODE_201_16 , NODE_374_8 , NODE_38_86 , NODE_486_20 , NODE_507_12 , NODE_52_12 , NODE_525_9 , NODE_558_28 , NODE_606_27 and NODE_98_38 ) were upregulated by at least 1.2-fold at 8 h, similar to expression of Crmapk that especially highly expressed at 8 h during the mycoparasitic process of C. chloroleuca 67-1 10 . To explore the roles and mechanisms of Crmapk in C. chloroleuca mycoparasitism, the gene was deleted using gene homologous recombination strategy, and the transcriptome analysis of ΔCrmapk mutant mycoparasitising S. sclerotiorum sclerotia was performed before 10 .…”

“…Combining with these results, we found that 38 interacting genes were differentially expressed (|Log 2 FC| ≥ 1 and p ≤ 0.05; Table 1 ), among which 15 genes ( NODE_15_2 , NODE_171_30 , NODE_176_9 , NODE_198_20 , NODE_2_38 , NODE_201_16 , NODE_374_8 , NODE_38_86 , NODE_486_20 , NODE_507_12 , NODE_52_12 , NODE_525_9 , NODE_558_28 , NODE_606_27 and NODE_98_38 ) were upregulated by at least 1.2-fold at 8 h, similar to expression of Crmapk that especially highly expressed at 8 h during the mycoparasitic process of C. chloroleuca 67-1 10 . To explore the roles and mechanisms of Crmapk in C. chloroleuca mycoparasitism, the gene was deleted using gene homologous recombination strategy, and the transcriptome analysis of ΔCrmapk mutant mycoparasitising S. sclerotiorum sclerotia was performed before 10 . In this study, it showed that when Crmapk was deficient 11 putative interacting genes from the Y2H library were differentially expressed (|Log 2 FC| ≥ 1 and p ≤ 0.05) based on the transcriptome profile of the C. chloroleuca mutant mycoparasitising sclerotia (Table 2 ), indicating that these genes might be involved in similar pathways to Crmapk .…”

“…In the previous study, we sequenced and analysed the transcriptome of the highly efficient C. chloroleuca strain 67-1 (formerly C. rosea 67-1) mycoparasitising the sclerotia of S. sclerotiorum 9 , from which we identified the mitogen-activated protein kinase (MAPK)-encoding gene Crmapk that is orthologous to a Fus3/Kss1 pathway member in Saccharomyces cerevisiae . Crmapk deficiency led to a reduced mycoparasitic ability to sclerotia and much lower biocontrol efficacy against soybean sclerotinia stem rot, indicating that Crmapk plays important roles in the biocontrol process of C. chloroleuca 10 . However, the molecular mechanism by which Crmapk regulates the mycoparasitism of C. chloroleuca remains unclear.…”

Deletion of all three MAP kinase genes results in severe defects in stress responses and pathogenesis in Fusarium graminearum

“…In F. graminearum, the phosphorylation of Mgv1 was inhibited by HopAI, an effector from Pseudomonas syringae (Zhang et al, 2017). Similar to their plant and animal counterparts, filamentous fungi have Nod-like immune receptors (NLRs) for signal recognition and activation of downstream targets (Uehling et al, 2017). F. graminearum has over 60 predicted NLR genes and some of them may be involved in recognizing MAMPS and fungal-bacterial interactions.…”

Early transcriptomic response of the mycoparasite Sphaerodes mycoparasitica to the mycotoxigenic Fusarium graminearum 3-ADON, the cause of Fusarium head blight

“…Several mycoparasitism-related differentially expressed genes (DEGs) have been reported. Signal transduction or signaling-related proteins, including G-proteins and mitogen-activated protein kinases (MAPKs), are involved in a series of processes, e.g., recognition or sensing the host or prey, (interspecific) communication, and attachment of parasitic appressorium-like structure(s) to the host (Omero et al 1999 ; Reithner et al 2007 ; Sun et al 2020 ; Yang 2017 ; Zeilinger et al 2005 ). In the S. mycoparasitica transcriptome, the expression levels of transcripts encoding G-proteins and MAPKs were higher during the interaction with the host than in controls (without host interactions) (data are not shown).…”