A highly conserved metalloprotease effector enhances virulence in the maize anthracnose fungus Colletotrichum graminicola

Abstract

Colletotrichum graminicola causes maize anthracnose, an agronomically important disease with a worldwide distribution. We have identified a fungalysin metalloprotease (Cgfl) with a role in virulence. Transcriptional profiling experiments and live cell imaging show that Cgfl is specifically expressed during the biotrophic stage of infection. To determine whether Cgfl has a role in virulence, we obtained null mutants lacking Cgfl and performed pathogenicity and live microscopy assays. The appressorium morphology of the null mutants is normal, but they exhibit delayed development during the infection process on maize leaves and roots, showing that Cgfl has a role in virulence. In vitro chitinase activity assays of leaves infected with wild-type and null mutant strains show that, in the absence of Cgfl, maize leaves exhibit increased chitinase activity. Phylogenetic analyses show that Cgfl is highly conserved in fungi. Similarity searches, phylogenetic analysis and transcriptional profiling show that C. graminicola encodes two LysM domain-containing homologues of Ecp6, suggesting that this fungus employs both Cgfl-mediated and LysM protein-mediated strategies to control chitin signalling.

Chitin and chitin-related compounds in plant–fungal interactions

“…In vitro chitinase activity assays of leaves infected with wild-type and null mutant strains show that, in the absence of Cgfl, maize leaves exhibit increased chitinase activity. Similarity searches, phylogenetic analysis, and transcriptional profiling revealed that C. graminicola encodes two LysM domain-containing homologs of Ecp6, suggesting that this fungus employs both Cgfl-mediated and LysM protein-mediated strategies to control chitin signalling (Sanz-Martín et al 2016). …”

Dual function of a secreted fungalysin metalloprotease in Ustilago maydis

“…Significant virulence reduction in the SG200∆umfly1 mutant also confirms the relevance of fungalysin in the U. maydis –maize interaction. Consistently, cgfl of C. graminicola was also found to be involved in virulence during maize infection (Sanz‐Martin et al ., ). The observed reduction of ZmChiA cleavage activity, as well as the reduced virulence both in the SG200∆umfly1 mutant and the active site mutant (SG200∆umfly1‐C mut ), indicates that the enzymatic activity of UmFly1 is required for the cleavage of maize ZmChiA, and consequently for U. maydis virulence.…”

“…Contrarily, in U. maydis , the deletion of umfly1 alone already results in a loss of ZmChiA cleavage activity and a reduced virulence of the fungus, indicating the importance of umfly1 for interaction with the host plant. Consistent with these results, cgfl of C. graminicola is also involved in virulence during maize infection (Sanz‐Martin et al ., ). Additionally, U. maydis also contains a single‐copy gene orthologous to fosep1 .…”

“…Consistent with this, Sanz‐Martin et al . () have shown that cgfl , a fungalysin‐encoding gene of C. graminicola , is specifically upregulated during the biotrophic phase of the infection process, and it is downregulated upon the switch from the biotrophic phase to the necrotrophic phase. These results suggest that the biotrophic pathogen U. maydis and the hemi‐biotroph C. graminicola both require fungalysin, particularly during biotrophic growth, when the fungus depends on suppression of host defense.…”

“…Recently, Sanz‐Martin et al . () showed that a fungalysin metalloprotease gene ( Cgfl ) in Colletotrichum graminicola is also involved in fungal virulence on maize, suggesting that chitinase‐cleavage activity of fungalysin is required for full virulence. Moreover, it has been shown that inhibition of fungalysin in Fusarium verticillioides by using a wheat hevein‐like peptide also inhibits the hyphal elongation, indicating that fungalysin plays a role in fungal development as well (Slavokhotova et al ., ).…”