{"title":"14-3-3 proteins inhibit autophagy by regulating SINAT-mediated proteolysis of ATG6 in Arabidopsis.","authors":"Ting Liu, Yuping Zheng, Shunkang Zhou, Yao Wang, Xue Lei, Lijuan Xie, Qingqi Lin, Changqing Chang, Shi Xiao, Rongliang Qiu, Hua Qi","doi":"10.1186/s12870-024-05854-3","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Autophagy is a conserved cellular process crucial for recycling cytoplasmic components and maintaining cellular homeostasis in eukaryotes. During autophagy, the formation of a protein complex involving AUTOPHAGY-RELATED PROTEIN 6 (ATG6) and phosphatidylinositol 3-kinase is pivotal for recruiting proteins involved in phagophore expansion. However, the intricate molecular mechanism regulating this protein complex in plants remains elusive.</p><p><strong>Results: </strong>Here, we aimed to unravel the molecular regulation of autophagy dynamics in Arabidopsis thaliana by investigating the involvement of the scaffold proteins 14-3-3λ and 14-3-3κ in regulating the proteolysis of ATG6. Phenotypic analyses revealed that 14-3-3λ and 14-3-3κ overexpression lines exhibited increased sensitivity to nutrient starvation, premature leaf senescence, and a decrease in starvation-induced autophagic vesicles, resembling the phenotypes of autophagy-defective mutants, suggesting the potential roles of 14-3-3 proteins in regulating autophagy in plants. Furthermore, our investigation unveiled the involvement of 14-3-3λ and 14-3-3κ in the RING finger E3 ligase SINAT1-mediated ubiquitination and destabilization of ATG6 in vivo. We also observed repressed turnover of ATG6 and translocation of GFP-ATG6 to mCherry-ATG8a-labelled punctate structures in the autophagy-defective mutant, which suggesting that ATG6 is probably a target of autophagy. Additionally, 14-3-3λ and 14-3-3κ interacted with Tumor necrosis factor Receptor Associated Factor 1a (TRAF1a) to promote the stability of TRAF1a in vivo under nutrient-rich conditions, suggesting a feedback regulation of autophagy. These findings demonstrate that 14-3-3λ and 14-3-3κ serve as scaffold proteins to regulate autophagy by facilitating the SINAT1-mediated proteolysis of ATG6, involving both direct and indirect mechanisms, in plants.</p><p><strong>Conclusions: </strong>14-3-3 proteins regulate autophagy by directly or indirectly binding to ATG6 and SINAT1 to promote ubiquitination and degradation of ATG6. 14-3-3 proteins are involved in modulating autophagy dynamics by facilitating SINAT1-mediated ubiquitination and degradation of ATG6.</p>","PeriodicalId":9198,"journal":{"name":"BMC Plant Biology","volume":"24 1","pages":"1148"},"PeriodicalIF":4.3000,"publicationDate":"2024-11-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11605875/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"BMC Plant Biology","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1186/s12870-024-05854-3","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"PLANT SCIENCES","Score":null,"Total":0}
引用次数: 0
Abstract
Background: Autophagy is a conserved cellular process crucial for recycling cytoplasmic components and maintaining cellular homeostasis in eukaryotes. During autophagy, the formation of a protein complex involving AUTOPHAGY-RELATED PROTEIN 6 (ATG6) and phosphatidylinositol 3-kinase is pivotal for recruiting proteins involved in phagophore expansion. However, the intricate molecular mechanism regulating this protein complex in plants remains elusive.
Results: Here, we aimed to unravel the molecular regulation of autophagy dynamics in Arabidopsis thaliana by investigating the involvement of the scaffold proteins 14-3-3λ and 14-3-3κ in regulating the proteolysis of ATG6. Phenotypic analyses revealed that 14-3-3λ and 14-3-3κ overexpression lines exhibited increased sensitivity to nutrient starvation, premature leaf senescence, and a decrease in starvation-induced autophagic vesicles, resembling the phenotypes of autophagy-defective mutants, suggesting the potential roles of 14-3-3 proteins in regulating autophagy in plants. Furthermore, our investigation unveiled the involvement of 14-3-3λ and 14-3-3κ in the RING finger E3 ligase SINAT1-mediated ubiquitination and destabilization of ATG6 in vivo. We also observed repressed turnover of ATG6 and translocation of GFP-ATG6 to mCherry-ATG8a-labelled punctate structures in the autophagy-defective mutant, which suggesting that ATG6 is probably a target of autophagy. Additionally, 14-3-3λ and 14-3-3κ interacted with Tumor necrosis factor Receptor Associated Factor 1a (TRAF1a) to promote the stability of TRAF1a in vivo under nutrient-rich conditions, suggesting a feedback regulation of autophagy. These findings demonstrate that 14-3-3λ and 14-3-3κ serve as scaffold proteins to regulate autophagy by facilitating the SINAT1-mediated proteolysis of ATG6, involving both direct and indirect mechanisms, in plants.
Conclusions: 14-3-3 proteins regulate autophagy by directly or indirectly binding to ATG6 and SINAT1 to promote ubiquitination and degradation of ATG6. 14-3-3 proteins are involved in modulating autophagy dynamics by facilitating SINAT1-mediated ubiquitination and degradation of ATG6.
背景:在真核生物中,自噬是一种保守的细胞过程,对细胞质成分的循环利用和维持细胞稳态至关重要。在自噬过程中,涉及自噬相关蛋白6 (autophagy - related protein 6, ATG6)和磷脂酰肌醇3-激酶的蛋白复合物的形成对于招募参与吞噬细胞扩张的蛋白至关重要。然而,在植物中调控这种蛋白复合物的复杂分子机制仍然是一个谜。结果:本研究旨在通过研究支架蛋白14-3-3λ和14-3-3κ参与ATG6蛋白水解的调控,揭示拟南芥自噬动力学的分子调控。表型分析显示,14-3-3λ和14-3-3κ过表达系对营养饥饿、叶片过早衰老和饥饿诱导的自噬囊泡减少的敏感性增加,与自噬缺陷突变体的表型相似,提示14-3-3蛋白在调节植物自噬中的潜在作用。此外,我们的研究揭示了14-3-3λ和14-3-3κ参与环指E3连接酶sinat1介导的体内ATG6的泛素化和不稳定。我们还观察到,在自噬缺陷突变体中,ATG6的转换受到抑制,GFP-ATG6易位到mcherry - atg8a标记的点状结构上,这表明ATG6可能是自噬的靶点。此外,14-3-3λ和14-3-3κ与肿瘤坏死因子受体相关因子1a (TRAF1a)相互作用,在营养丰富的条件下促进TRAF1a在体内的稳定性,提示自噬的反馈调节。这些发现表明14-3-3λ和14-3-3κ作为支架蛋白,通过促进sinat1介导的ATG6蛋白水解来调节植物自噬,包括直接和间接机制。结论:14-3-3蛋白通过直接或间接结合ATG6和SINAT1促进ATG6的泛素化和降解来调节自噬。14-3-3蛋白通过促进sinat1介导的泛素化和ATG6降解参与调节自噬动力学。
期刊介绍:
BMC Plant Biology is an open access, peer-reviewed journal that considers articles on all aspects of plant biology, including molecular, cellular, tissue, organ and whole organism research.