Mitogen-activated protein kinase (MAPK) signaling pathways are evolutionarily conserved in eukaryotes and modulate responses to both internal and external stimuli. Pmk1 and Mps MAPK pathways regulate stress tolerance, vegetative growth and cell wall integrity in Saccharomyces cerevisiae and Pyricularia oryzae. Here, we deployed genetic and cell biology strategies to investigate the roles of the orthologs of Pmk1 and Mps1 in Sclerotiophoma versabilis (herein referred to as SvPmk1 and SvMps1, respectively). Our results showed that SvPmk1 and SvMps1 are involved in hyphal development, asexual reproduction and pathogenesis in S. versabilis. We found that ∆Svpmk1 and ∆Svmps1 mutants have significantly reduced vegetative growths on PDA supplemented with osmotic stress-inducing agents, compared to the wild type, with ∆Svpmps1 being hypersensitive to hydrogen peroxide. The two mutants failed to produce pycnidia and have reduced pathogenicity on Pseudostellaria heterophylla. Unlike SvPmk1, SvMps1 was found to be indispensable for the fungal cell wall integrity. Confocal microscopic analyses revealed that SvPmk1 and SvMps1 are ubiquitously expressed in the cytosol and nucleus. Taken together, we demonstrate here that SvPmk1 and SvMps1 play critical roles in the stress resistance, development and pathogenesis of S. versabilis.

Ascochyta versabilis is the fungal pathogen that causes the severe leaf spot disease of Pseudostellaria heterophylla (Miq.) Pax, a vital Chinese herbal plant. Here we deployed PacBio SMRT long-read sequencing technology to generate a near-complete genome assembly for the A. versabilis KC1 strain and obtained a total of 9.80 Gb raw reads. These reads were processed into a 41.05 Mb genome assembly containing 95 contigs with N50 of 1.70 Mb and a maximum length of 3.93 Mb. A total of 10,457 gene models, of which 1,004 encode putatively secreted proteins, were identified in the genome. This high-quality genome assembly and gene annotation resource reported here will facilitate the institution of functional genetic studies aimed at providing a better insight into the infection mechanisms of A. versabilis to support the development of effective control strategies for leaf spot disease of P. heterophylla.

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Protein kinases and phosphatases catalyze the phosphorylation and dephosphorylation of their protein substrates, respectively, and these are important mechanisms in cellular signal transduction. The rice blast fungus Magnaporthe oryzae possesses 6 protein phosphatases of type 2C class, including MoPtc1, 2, 5, 6, 7 and 8. However, only very little is known about the roles of these phosphatases in filamentous fungi. Here in, we deployed genetics and molecular biology techniques to identify, characterize and establish the roles of MoPtc5 and MoPtc7 in M. oryzae development and pathogenicity. We found that during pathogen-host interaction, MoPTC7 is differentially expressed. Double deletion of MoPTC7 and MoPTC5 suppressed the fungal vegetative growth, altered its cell wall integrity and reduced its virulence. The two genes were found indispensable for stress tolerance in the phytopathogen. We also demonstrated that disruption of any of the two genes highly affected appressorium turgor generation and Mps1 and Osm1 phosphorylation levels. Lastly, we demonstrated that both MoPtc5 and MoPtc7 are localized to mitochondria of different cellular compartments in the blast fungus. Taken together, our study revealed synergistic coordination of M. oryzae development and pathogenesis by the type 2C protein phosphatases.

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