The antifungal protein afpb induces regulated cell death in its parental fungus penicillium digitatum
Bugeda, Adrià (Centre de Recerca en Agrigenòmica)
Garrigues, Sandra (Consejo Superior de Investigaciones Científicas (Espanya). Instituto de Agroquimica y Tecnologia de Alimentos)
Gandía, Mónica (Consejo Superior de Investigaciones Científicas (Espanya). Instituto de Agroquimica y Tecnologia de Alimentos)
Manzanares, Paloma (Consejo Superior de Investigaciones Científicas (Espanya). Instituto de Agroquimica y Tecnologia de Alimentos)
Marcos, Jose F. (Consejo Superior de Investigaciones Científicas (Espanya). Instituto de Agroquimica y Tecnologia de Alimentos)
Coca López, María (Centre de Recerca en Agrigenòmica)

Data:	2020
Resum:	Filamentous fungi produce small cysteine-rich proteins with potent, specific antifungal activity, offering the potential to fight fungal infections that severely threaten human health and food safety and security. The genome of the citrus postharvest fungal pathogen Penicillium digitatum encodes one of these antifungal proteins, namely AfpB. Biotechnologically produced AfpB inhibited the growth of major pathogenic fungi at minimal concentrations, surprisingly including its parental fungus, and conferred protection to crop plants against fungal infections. This study reports an in-depth characterization of the AfpB mechanism of action, showing that it is a cell-penetrating protein that triggers a regulated cell death program in the target fungus. We prove the importance of AfpB interaction with the fungal cell wall to exert its killing activity, for which protein mannosylation is required. We also show that the potent activity of AfpB correlates with its rapid and efficient uptake by fungal cells through an energy-dependent process. Once internalized, AfpB induces a transcriptional reprogramming signaled by reactive oxygen species that ends in cell death. Our data show that AfpB activates a self-injury program, suggesting that this protein has a biological function in the parental fungus beyond defense against competitors, presumably more related to regulation of the fungal population. Our results demonstrate that this protein is a potent antifungal that acts through various targets to kill fungal cells through a regulated process, making AfpB a promising compound for the development of novel biofungicides with multiple fields of application in crop and postharvest protection, food preservation, and medical therapies. IMPORTANCE Disease-causing fungi pose a serious threat to human health and food safety and security. The limited number of licensed antifungals, together with the emergence of pathogenic fungi with multiple resistance to available antifungals, represents a serious challenge for medicine and agriculture. Therefore, there is an urgent need for new compounds with high fungal specificity and novel antifungal mechanisms. Antifungal proteins in general, and AfpB from Penicillium digitatum in particular, are promising molecules for the development of novel antifungals. This study on AfpB's mode of action demonstrates its potent, specific fungicidal activity through the interaction with multiple targets, presumably reducing the risk of evolving fungal resistance, and through a regulated cell death process, uncovering this protein as an excellent candidate for a novel biofungicide. The in-depth knowledge on AfpB mechanistic function presented in this work is important to guide its possible future clinical and agricultural applications.
Ajuts:	Ministerio de Ciencia e Innovación BIO2015-68790-C2-1-R Ministerio de Ciencia e Innovación BIO2015-68790-C2-2-R Ministerio de Ciencia e Innovación RTI2018-101115B-C21 Ministerio de Ciencia e Innovación RTI2018-101115B-C22 Ministerio de Ciencia e Innovación SEV-2015-0533 Ministerio de Educación, Cultura y Deporte FPU13-04584
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	Antifungal proteins ; AFP ; Phytopathogens ; Penicillium digitatum ; Regulated cell death ; Cysteine-rich proteins ; Cell-penetrating protein ; Plant protection ; Fungal infection
Publicat a:	mSphere, Vol. 5, Issue 4 (August 2020) , art. e00595-20, ISSN 2379-5042

DOI: 10.1128/MSPHERE.00595-20
PMID: 32848004

15 p, 2.6 MB

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