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

Yufeng Liu,Yinggao Liu,Chunlin Yang,Qian Zeng,Fred O Asiegbu,Lijuan Liu,Shan Han,Shuai Yang

doi:10.3390/f12091268

Yufeng Liu, Yinggao Liu + Show 6 more

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https://doi.org/10.3390/f12091268

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Journal: Forests	Publication Date: Sep 17, 2021
Citations: 6	License type: CC BY 4.0

Affiliation: University of Helsinki, Sichuan Agricultural University

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.

Highlights

Forest trees are constantly faced with biotic and abiotic stresses throughout their life cycle, and have evolved a variety of mechanisms to protect themselves against these adverse environmental challenges [1]
The full-length cDNA sequences of four PlTLP genes from P. likiangensis were cloned based on the transcriptome library of P. likiangensis
Four members of the PR5 family were successfully isolated and identified from P. likiangensis. qPCR analysis showed that the expression levels of four PlTLP genes were significantly up-regulated by L. piceae invasion

Summary

Introduction

Forest trees are constantly faced with biotic and abiotic stresses throughout their life cycle, and have evolved a variety of mechanisms to protect themselves against these adverse environmental challenges [1]. PR proteins are generally classified into 17 different families based on their physicochemical properties and biological functions, and most of them have antifungal activity [5,6]. Most TLP proteins contain a thaumatin signature (G-x-[GF]-x-C-xT-[GA]-D-C-x(1,2)-[GQ]-x(2,3)-C) with a molecular mass in the range of 21–26 kDa, and 16 cysteine residues that form eight disulfide bonds. A small number of TLP proteins have a molecular weight of approximately 16–17 kDa and only contain 10 conserved cysteine residues that form five disulfide bonds [10,11]. These disulfide bonds contribute to maintaining the stability of the protein structure, and strengthening the resistance of TLP proteins to protease degradation, extreme temperature, and pH conditions [12,13]

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