Mining oomycete proteomes for metalloproteases leads to identification of candidate virulence factors in Phytophthorainfestans

Abstract

Pathogens deploy a wide range of pathogenicity factors, including a plethora of proteases, to modify host tissue or manipulate host defences. Metalloproteases (MPs) have been implicated in virulence in several animal and plant pathogens. Here we investigated the repertoire of MPs in 46 stramenopile species including 37 oomycetes, 5 diatoms, and 4 brown algae. Screening their complete proteomes using hidden Markov models (HMMs) trained for MP detection resulted in over 4,000 MPs, with most species having between 65 and 100 putative MPs. Classification in clans and families according to the MEROPS database showed a highly diverse MP repertoire in each species. Analyses of domain composition, orthologous groups, distribution, and abundance within the stramenopile lineage revealed a few oomycete‐specific MPs and MPs potentially related to lifestyle. In‐depth analyses of MPs in the plant pathogen Phytophthora infestans revealed 91 MPs, divided over 21 protein families, including 25 MPs with a predicted signal peptide or signal anchor. Expression profiling showed different patterns of MP gene expression during pre‐infection and infection stages. When expressed in leaves of Nicotiana benthamiana, 12 MPs changed the sizes of lesions caused by inoculation with P. infestans; with 9 MPs the lesions were larger, suggesting a positive effect on the virulence of P. infestans, while 3 MPs had a negative effect, resulting in smaller lesions. To the best of our knowledge, this is the first systematic inventory of MPs in oomycetes and the first study pinpointing MPs as potential pathogenicity factors in Phytophthora.

RNA silencing proteins and small RNAs in oomycete plant pathogens and biocontrol agents

“…Several metalloproteases have been characterized as virulence factors for Ph. infestans (Schoina et al, 2021), and the potential for targeting enzymes of this class by Py. periplocum milRNAs suggests the possibility for this oomycete to use cross-kingdom RNA silencing as a component of its biocontrol action.…”

The root pathogen Aphanomyces euteiches secretes modular proteases in pea apoplast during host infection

“…Recent evidence has shown that proteases from oomycetes might also have a role to play during plant invasion. Expression profile of in-silico identified metalloprotease from Phytophthora infestans pinpointed a dozen of enzymes that potentially affect virulence of the pathogen ( Schoina et al., 2021 ). Knockout-mutants of two secreted cysteine proteases from Phytophthora parasitica (PpCys44, 45) present a reduced virulence during Nicotiana benthamiana infection, while overexpression of both proteases in the plant apoplast triggers cell death ( Zhang et al., 2020 ).…”

The dynamic organization of the actin and tubulin cytoskeleton in Phytophthora pathogens

“…The vegetative propagules of P. infestans are sporangia that germinate directly or indirectly, depending on the ambient temperature. At temperatures lower than 15 °C the sporangia cleave and release motile zoospores, while at higher temperatures the sporangia can germinate directly (Judelson & Blanco 2005, Schoina & Govers 2015. When encountering a suitable environment, like a leaf surface, the hyphal germlings emerging from sporangia or from encysted zoospores develop an appressorium at the tip, and subsequently a penetration peg is formed that pierces the plant epidermis.…”

“…When encountering a suitable environment, like a leaf surface, the hyphal germlings emerging from sporangia or from encysted zoospores develop an appressorium at the tip, and subsequently a penetration peg is formed that pierces the plant epidermis. After the pathogen has gained access to the plant, the hyphae grow intercellular in the mesophyll occasionally forming digit-like structures called haustoria that penetrate plant cells (Judelson & Blanco 2005, Schoina & Govers 2015. Contrary to fungal hyphae, the hyphae of oomycetes lack septa or cross walls and are therefore referred to as aseptate or coenocytic.…”

“…All MAPs are clustered according to predicted function and/or subcellular localization (grey rows). Expression profiles are based on RNAseq data of P. infestans strain 88069(Schoina et al 2021). Abbreviations: myc mycelium, spor sporangia, zsp zoospores, gc germinating cysts.…”