Abstract
Fungi are the most prevalent plant pathogens, causing annually important damages. To infect and colonize their hosts, they secrete effectors including hydrolytic enzymes able to kill and macerate plant tissues. These secreted proteins are transported from the Endoplasmic Reticulum and the Golgi apparatus to the extracellular space through intracellular vesicles. In pathogenic fungi, intracellular vesicles were described but their biogenesis and their role in virulence remain unclear. In this study, we report the essential role of clathrin heavy chain (CHC) in the pathogenicity of Botrytis cinerea, the agent of gray mold disease. To investigate the importance of this protein involved in coat vesicles formation in eukaryotic cells, a T-DNA insertional mutant reduced in the expression of the CHC-encoding gene, and a mutant expressing a dominant-negative form of CHC were studied. Both mutants were strongly affected in pathogenicity. Characterization of the mutants revealed altered infection cushions and an important defect in protein secretion. This study demonstrates the essential role of clathrin in the infectious process of a plant pathogenic fungus and more particularly its role in virulence factors delivery.
Highlights
Filamentous fungi constitute the major group of crop pathogenic microbes that reduce the yield and quality of agricultural production
Random mutagenesis leading to the Abbreviations: clathrin heavy chain (CHC), Clathrin Heavy Chain; Agrobacterium tumefaciensmediated transformation (ATMT), Agrobacterium tumefaciens-mediated transformation; plasma membrane (PM), Plasma Membrane; trans-Golgi network (TGN), Trans Golgi Network; clathrin coat vesicles (CCV), Clathrin Coat Vesicles; clathrin light chains (CLC), Clathrin Light Chain; clathrin-mediated endocytosis (CME), Clathrin Mediated Endocytosis; reactive oxygen species (ROS), Reactive Oxygen Species; signal peptide (SP), Signal Peptide; carbohydrate-active enzymes (CAZy), Carbohydrate Active Enzymes; PCWDE, Plant Cell Wall Degrading Enzymes; fungal cell wall enzymes (FCWE) Fungal Cell Wall Enzymes; cell death inducing proteins (CDIPs), Cell Death-Inducing Proteins
Southern-blot analysis of this mutant showed a single insertion of the bacterium T-DNA in the fungal genome (Supplementary Figures 1A,C) and a T-DNA rescue approach localized this insertion within the promoter region of the CHC-encoding gene (Bcchc–Bcin01g09940), 297 pb upstream of the ATG (Figure 1A)
Summary
Filamentous fungi constitute the major group of crop pathogenic microbes that reduce the yield and quality of agricultural production. New environmentally friendly crop protection methods are needed to limit the presence of residues in soil and the continual emergence of new races of fungi able to overcome chemical treatments (Shang et al, 2019). Within this context, a better understanding of the infectious process developed by plant pathogenic fungi becomes a prerequisite to develop new and alternative solutions. Random mutagenesis leading to the Abbreviations: CHC, Clathrin Heavy Chain; ATMT, Agrobacterium tumefaciens-mediated transformation; PM, Plasma Membrane; TGN, Trans Golgi Network; CCV, Clathrin Coat Vesicles; CLC, Clathrin Light Chain; CME, Clathrin Mediated Endocytosis; ROS, Reactive Oxygen Species; SP, Signal Peptide; CAZy, Carbohydrate Active Enzymes; PCWDE, Plant Cell Wall Degrading Enzymes; FCWE Fungal Cell Wall Enzymes; CDIPs, Cell Death-Inducing Proteins
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