Functional Analysis of a Tannic-Acid-Inducible and Hypoviral-Regulated Small Heat-Shock Protein Hsp24 from the Chestnut Blight Fungus Cryphonectria parasitica

Abstract

A small heat-shock protein gene, CpHsp24, of Cryphonectria parasitica was selected based on its expression pattern, which showed that it was tannic acid inducible and that its induction was severely hampered by a hypovirus. The predicted protein sequence of CpHsp24 consisted of a hallmark α-crystalline domain flanked by a variable N-terminal and a short C-terminal region. Disruption of CpHsp24 resulted in a slow growth rate under standard growth conditions. The CpHsp24-null mutant showed enhanced sensitivity to heat shock, which was consistent with Northern and Western analyses displaying the heat-shock induction of the CpHsp24 gene and protein, respectively. Virulence tests on the excised bark revealed a severe decrease in the necrotic area of the CpHsp24-null mutant. When the hypovirus was transferred, virus-containing CpHsp24-null progeny displayed severely retarded growth patterns with hypovirulent characteristics of reduced pigmentation and sporulation. Because the tannic-acid-inducible and hypoviral-suppressible expression and the severely impaired virulence are also characteristics of the laccase3 gene (lac3), lac3 expression in the CpHsp24-null mutant was also examined. The resulting lac3 induction was severely affected in the CpHsp24-null mutant, suggesting that CpHsp24 is important for lac3 induction and that CpHsp24 may act as a molecular chaperone for the lac3 protein.