PR proteins in Moniliophthora perniciosa: genomic and functional analysis of PR-1 and PR-5 (thaumatin - like) proteins

Author(s)

Renata Moro Baroni

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2016-02-18

Examining board members

Gonçalo Amarante Guimarães Pereira; Celso Eduardo Benedetti; Jörg Kobarg; Ronaldo José Durigan Dalio; Gonçalo Apolinario de Souza Filho

• Advisor: Jorge Mauricio Costa Mondego; Gonçalo Amarante Guimarães Pereira

Abstract

The basidiomycete fungus Moniliophthora perniciosa is the causative agent of Witches¿ Broom disease (WBD) in cacao (Theobroma cacao). M. perniciosa displays a hemibiotrophic lifestyle with an extended biotrophic stage, an uncommon trait among phytopathogens. The genome of M. perniciosa, enhanced by data originated in RNA-seq, allowed annotation of genes encoding Pathogenicity Related Protein (PRs). The PRs are a group of 17 functionally heterogeneous protein families, among which we can highlight the PR-1 and the PR-5 (Thaumatin-like). Thaumatin-like proteins (TLPs) are considered fungal inhibitors in plants. However, studies in basidiomycete fungal show their involvement in cellular remodeling during mushrooms formation. In this work, we have identified that M. perniciosa fungus is the species with greatest number of Thaumatin-like, with 13 genes. RNA-seq annotation and qPCR experiments revealed that 4 MpTLPs are expressed during the dry broom stage of the disease, at which the cacao is susceptible to opportunistic fungus, indicating that these proteins are involved in the fight against these microrganisms. In addition, we hypothesize the participation of TLPs in basidiocarp formation and in the fungal protection against plant defense mechanisms triggered by cell walls polysaccharides. Among the fungal proteins which may be significant for establish the disease are proteins with conserved domains SCP/TAPS, also known as PR-1. Several articles have shown the importance of these proteins in the immune response system not only in plants but also in various organisms, from yeast to humans. Furthermore, SCP/TAPS proteins also have the ability to inhibit the action of hydrophobic plant molecules with antifungal potential. We identified in the genome of M. perniciosa 11 genes (MpPR-1a ¿ MpPR1-k) encoding proteins belonging to the superfamily SCP/TAPS. Those are differentially expressed throughout the development of the fungus and some of them are preferentially or exclusively expressed in plant during the progression of the witch's broom disease. Functional assays using complementation of mutant yeasts depleted of PR-1 genes (PRY) and lipid-protein binding experiments, shown that MpPR-1s are able to export and bind cholesterol and palmitate, and also to help fungal detoxification against the effects of the plant glycosteroid tomatin. We believe that this work can contribute to a better understanding of the mechanisms about the functions of MpPR1s and MpTLPs on WBD. In addition, to contributing the efforts being undertaken in the Witches¿ broom Project, transforming the knowledge gained in possible targets to control WBD. Besides of adding new information that converted into knowledge can generate potential targets for the dream control of Witches¿ Broom disease (AU)