异源过表达蒺藜苜蓿<i>MtATG7</i>基因促进拟南芥抵抗碳氮饥饿胁迫

朱晓洁; 王贤杰; 杨明康; 黄巍; 陈亮

doi:10.7671/j.issn.1001-411X.202207033

异源过表达蒺藜苜蓿MtATG7基因促进拟南芥抵抗碳氮饥饿胁迫

Heterologous overexpression of MtATG7 gene from Medicago truncatula promotes resistance to carbon and nitrogen starvation in Arabidopsis thaliana

摘要

摘要:
目的真核生物通过保守的自噬途径降解错误折叠的蛋白质或受损细胞器，实现对营养物质的循环再利用。本文旨在解析苜蓿自噬基因在植物应对营养胁迫中发挥的功能，为指导苜蓿的选育和改良提供参考。
方法以自噬途径的关键限速基因ATG7(Autophagy-related gene 7)为切入点，分析ATG7氨基酸序列在不同植物中的相似性。利用蒺藜苜蓿Medicago truncatula的MtATG7基因过表达载体转化拟南芥植株，获得异源过表达植株35S::MtATG7和互补拟南芥突变体的atg7/35S::MtATG7植株，并对其抗胁迫表型和自噬活性进行分析。
结果在碳饥饿条件下，atg7/35S::MtATG7能够挽救atg7突变体叶片早衰的表型；恢复到正常生长条件以后，35S::MtATG7和atg7/35S::MtATG7植株存活率和野生型相比都显著提高。GFP-ATG8e的剪切试验表明，atg7/35S::MtATG7能够恢复atg7突变体的自噬降解活性。在氮饥饿条件下过表达MtATG7同样能够减缓拟南芥叶片的衰老速度。
结论异源过表达MtATG7能增强拟南芥碳氮饥饿抗性的生物学功能，本研究为进一步利用MtATG7基因改良苜蓿农艺性状提供了理论依据。

Abstract:
Objective Eukaryotes use autophagy pathway to recycle nutrients by degrading misfolded proteins or damaged organelles. This study aims to analyze the function of Medicago autophagy genes in plant response to nutritional stress, and provide a reference for the breeding and improvement of Medicago.
Method We focused on the ATG7 gene (Autophagy-related gene 7), which serves as the rate-limiting gene in the autophagy pathway, and investigated the similarities of ATG7 amino acid sequences among different plant species. The overexpression vector of Medicago truncatula MtATG7 gene was transformed to Arabidopsis thaliana, to generate the heterologous overexpression lines 35S::MtATG7 and the complementary lines atg7/35S::MtATG7. The plant resistance to stress and autophagic activity were analyzed.
Result Under carbon starvation, atg7/35S::MtATG7 rescued the phenotype of premature leaf senescence of atg7 mutant, and the survival rates of 35S::MtATG7 and atg7/35S::MtATG7 plants were significantly higher than that of wildtype. GFP-ATG8e cleavage assay suggested that atg7/35S::MtATG7 restored the deficiency of autophagic degradation activity in atg7 mutants. Under nitrogen starvation, overexpression of MtATG7 also slowed down the senescence rate of A. thaliana leaves.
Conclusion Heterologous overexpression of MtATG7 can enhance the resistance of A. thaliana under carbon and nitrogen starvation, which provides a theoretical basis for further improving Medicago agronomic traits by using MtATG7 gene.

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