Molecular Identification and Antifungal Properties of Four Thaumatin-like Proteins in Spruce (Picea likiangensis)

Abstract

Thaumatin-like proteins (TLPs) are involved in the plant defense response against pathogens, and most of them exhibit antifungal activity. However, the role of TLPs in pathogen-induced defense responses in spruce is not fully understood. In this study, four TLP genes encoding thaumatin-like protein, designated as PlTLP1–4, were isolated and identified from Picea likiangensis needles. Sequence analysis showed that PlTLP1, PlTLP3, and PlTLP4 contained 16 conserved cysteine residues, while PlTLP2 had only 10 conserved cysteine residues. qPCR analysis showed that PlTLPs were expressed in all tissues tested, PlTLP1, PlTLP3, and PlTLP4 had the highest expression levels in young fruits, while PlTLP2 had the highest expression levels in roots. In addition, the expression levels of four PlTLPs were significantly upregulated during infection by Lophodermium piceae. Four recombinant PlTLPs expressed in Escherichia coli exhibited obvious β-1,3-glucanase activity. The antifungal activity assay showed that four recombinant PlTLPs had significant inhibitory effects on the mycelial growth of L. piceae, Fusarium proliferatum, Botrytis cinerea, and Roussoella doimaesalongensis. Microscopic observation revealed that the recombinant PlTLP1–4 induced the morphological changes of the mycelia of L. piceae, and the recombinant PlTLP2 and PlTLP3 induced the morphological changes of the mycelia of F. proliferatum and R. doimaesalongensis, while all the recombinant PlTLPs had no obvious negative effect on the morphology of B. cinerea mycelium. These results suggest that PlTLP genes may play an important role in the defense response of P. likiangensis against L. piceae invasion.