Centre for Cell & Developmental Biology and State Key Laboratory of Agrobiotechnology, School of Life Sciences, The Chinese University of Hong Kong, Hong Kong, China.
Guangdong Provincial Key Laboratory of Biotechnology for Plant Development, School of Life Sciences, South China Normal University, Guangzhou, China.
Nat Commun. 2023 Mar 30;14(1):1768. doi: 10.1038/s41467-023-37185-6.
The energy sensor AMP-activated protein kinase (AMPK) can activate autophagy when cellular energy production becomes compromised. However, the degree to which nutrient sensing impinges on the autophagosome closure remains unknown. Here, we provide the mechanism underlying a plant unique protein FREE1, upon autophagy-induced SnRK1α1-mediated phosphorylation, functions as a linkage between ATG conjugation system and ESCRT machinery to regulate the autophagosome closure upon nutrient deprivation. Using high-resolution microscopy, 3D-electron tomography, and protease protection assay, we showed that unclosed autophagosomes accumulated in free1 mutants. Proteomic, cellular and biochemical analysis revealed the mechanistic connection between FREE1 and the ATG conjugation system/ESCRT-III complex in regulating autophagosome closure. Mass spectrometry analysis showed that the evolutionary conserved plant energy sensor SnRK1α1 phosphorylates FREE1 and recruits it to the autophagosomes to promote closure. Mutagenesis of the phosphorylation site on FREE1 caused the autophagosome closure failure. Our findings unveil how cellular energy sensing pathways regulate autophagosome closure to maintain cellular homeostasis.
当细胞能量产生受到损害时，能量传感器 AMP 激活蛋白激酶 (AMPK) 可以激活自噬。然而，营养感应对自噬体闭合的影响程度尚不清楚。在这里，我们提供了一种植物特有的蛋白质 FREE1 的作用机制，在自噬诱导的 SnRK1α1 介导的磷酸化作用下，FREE1 作为 ATG 连接系统和 ESCRT 机制之间的连接物，在营养缺乏时调节自噬体闭合。使用高分辨率显微镜、3D 电子断层扫描和蛋白酶保护分析，我们表明在 free1 突变体中积累了未闭合的自噬体。蛋白质组学、细胞和生化分析揭示了 FREE1 与 ATG 连接系统/ESCRT-III 复合物在调节自噬体闭合中的机制联系。质谱分析表明，进化保守的植物能量传感器 SnRK1α1 磷酸化 FREE1 并将其募集到自噬体上以促进闭合。FREE1 上磷酸化位点的突变导致自噬体闭合失败。我们的发现揭示了细胞能量感应途径如何调节自噬体闭合以维持细胞内稳态。
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