The Myosin Motor Domain-Containing Chitin Synthases Are Involved in Cell Wall Integrity and Sensitivity to Antifungal Proteins in Penicillium digitatum

Abstract

Penicillium digitatum is the main postharvest pathogen of citrus fruit and is responsible for important economic losses in spite of the massive use of fungicides. The fungal cell wall (CW) and its specific component chitin are potential targets for the development of new antifungal molecules. Among these are the antifungal peptides and proteins that specifically interact with fungal CW. Chitin is synthesized by a complex family of chitin synthases (Chs), classified into up to eight classes within three divisions. Previously, we obtained and characterized a mutant of P. digitatum in the class VII gene (ΔchsVII), which contains a short myosin motor-like domain (MMD). In this report, we extend our previous studies to the characterization of mutants in chsII and in the gene coding for the other MMD-Chs (chsV), and study the role of chitin synthases in the sensitivity of P. digitatum to the self-antifungal protein AfpB, and to AfpA obtained from P. expansum. The ΔchsII mutant showed no significant phenotypic and virulence differences with the wild type strain, except in the production and morphology of the conidia. In contrast, mutants in chsV showed a more dramatic phenotype than the previous ΔchsVII, with reduced growth and conidial production, increased chitin content, changes in mycelial morphology and a decrease in virulence to citrus fruit. Mutants in chsVII were specifically more tolerant than the wild type to nikkomycin Z, an antifungal inhibitor of chitin biosynthesis. Treatment of P. digitatum with its own antifungal protein AfpB resulted in an overall reduction in the expression of the chitin synthase genes. The mutants corresponding to MMD chitin synthases exhibited differential sensitivity to the antifungal proteins AfpA and AfpB, ΔchsVII being more susceptible than its parental strain and ΔchsV being slightly more tolerant despite its reduced growth in liquid broth. Taking these results together, we conclude that the MMD-containing chitin synthases affect cell wall integrity and sensitivity to antifungal proteins in P. digitatum.

Effects of Peptide C12-OOWW-NH2 on Transcriptome and Cell Wall of the Postharvest Fungal Pathogen Penicillium digitatum

“…In this study, the fewer and weaker blue fluorescence septa were observed under C 12 O 3 TR stress (Figure 5), indicating that the chitin content in the cell wall was influenced by C 12 O 3 TR treatment. This result was similar with the effect of antifungal proteins on cell wall chitin (Gandía et al, 2019). It was reported that the changes of chitin content was related to the activity of related enzymes, such as β-1, 3-glucan synthase and chitinase (OuYang et al, 2019).…”

Unveiling the Role Displayed by Penicillium digitatum PdMut3 Transcription Factor in Pathogen–Fruit Interaction

“…These effects have already been described in other cases where the cell wall was clearly damaged, for instance, in PdCrz1 mutants [26]. This result was also similar to the one observed with the addition of antifungal proteins on cell wall chitin [34]. The cell membrane plays a relevant role in maintaining cell viability because it represents a barrier by separating the cell from its environment, and is also a means of exchange of substances and energy between the cell and the surrounding environment.…”

Physiological and Proteomic Analysis of Penicillium digitatum in Response to X33 Antifungal Extract Treatment

“…The cell wall is the first barrier of filamentous fungi resist environmental pressures and is crucial for maintaining the normal growth of fungal ( Gandía et al, 2019 ). In the current study, chitin synthase (gi| 859266290 ), endoglucanase (gi| 859262625 ), 1,3-β-glucanosyltransferase (gi| 859262943 ), and β-glucosidase (gi| 859260053 ), which are important cell wall-related proteins, were overexpressed.…”