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.

中文翻译：

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云杉（Picea likiangensis）中四种奇异果甜蛋白样蛋白的分子鉴定和抗真菌特性

奇异果甜蛋白样蛋白 (TLPs) 参与植物对病原体的防御反应，其中大多数表现出抗真菌活性。然而，TLPs 在云杉病原体诱导的防御反应中的作用尚不完全清楚。在这项研究中，从丽江云杉针叶中分离并鉴定了四个编码奇异果甜蛋白样蛋白的TLP基因，命名为PlTLP1-4。序列分析显示PlTLP1、PlTLP3和PlTLP4含有16个保守的半胱氨酸残基，而PlTLP2只有10个保守的半胱氨酸残基。qPCR 分析表明PlTLPs在所有测试组织中均有表达，PlTLP1、PlTLP3和PlTLP4在幼果中的表达水平最高，而PlTLP2在根中的表达水平最高。此外，四种PlTLP的表达水平在云杉科感染期间显着上调。在大肠杆菌中表达的四种重组PlTLP表现出明显的 β-1,3-葡聚糖酶活性。抗真菌活性试验表明，4种重组PlTLPs对云杉、增殖镰刀菌、灰葡萄孢和灰霉病菌的菌丝体生长有显着抑制作用。Roussoella doimaesalongensis。显微镜观察显示，重组PlTLP1-4诱导的菌丝体的形态变化L.云杉，重组PlTLP2和PlTLP3诱导的菌丝体的形态变化F. proliferatum和R. doimaesalongensis，而所有的重组PlTLPs没有对B. cinerea菌丝体的形态有明显的负面影响。这些结果表明，PlTLP基因可能在丽江松对云杉入侵的防御反应中起重要作用。