Interaction between hypoviral-regulated fungal virulence factor laccase3 and small heat shock protein Hsp24 from the chestnut blight fungus Cryphonectria parasitica.

Laccase3 is an important virulence factor of the fungus Cryphonectria parasitica. Laccase3 gene (lac3) transcription is induced by tannic acid, a group of phenolic compounds found in chestnut trees, and its induction is regulated by the hypovirus CHV1 infection. CpHsp24, a small heat shock protein gene of C. parasitica, plays a determinative role in stress adaptation and pathogen virulence. Having uncovered in our previous study that transcriptional regulation of the CpHsp24 gene in response to tannic acid supplementation and CHV1 infection was similar to that of the lac3, and that conserved phenotypic changes of reduced virulence were observed in mutants of both genes, we inferred that both genes were implicated in a common pathway. Building on this finding, in this paper we examined whether the CpHsp24 protein (CpHSP24) was a molecular chaperone for the lac3 protein (LAC3). Our pull-down experiment indicated that the protein products of the two genes directly interacted with each other. Heterologous co-expression of CpHsp24 and lac3 genes using Saccharomyces cerevisiae resulted in more laccase activity in the cotransformant than in a parental lac3-expresssing yeast strain. These findings suggest that CpHSP24 is, in fact, a molecular chaperone for the LAC3, which is critical component of fungal pathogenesis.