Oxidative stress underlies a number of pathological conditions, including cancer, neurodegeneration, and aging. Antioxidant-rich foods help maintain cellular redox homeostasis and mitigate oxidative stress, but the underlying mechanisms are not clear. For example, sulforaphane (SFN), an electrophilic compound that is enriched in cruciferous vegetables such as broccoli, is a potent inducer of cellular antioxidant responses. NFE2L2/NRF2 (nuclear factor, erythroid 2 like 2), a transcriptional factor that controls the expression of multiple detoxifying enzymes through antioxidant response elements (AREs), is a proposed target of SFN. NFE2L2/NRF2 is a target gene of TFEB (transcription factor EB), a master regulator of autophagic and lysosomal functions, which we show here to be potently activated by SFN. SFN induces TFEB nuclear translocation via a Ca²⁺-dependent but MTOR (mechanistic target of rapamycin kinase)-independent mechanism through a moderate increase in reactive oxygen species (ROS). Activated TFEB then boosts the expression of genes required for autophagosome and lysosome biogenesis, which are known to facilitate the clearance of damaged mitochondria. Notably, TFEB activity is required for SFN-induced protection against both acute oxidant bursts and chronic oxidative stress. Hence, by simultaneously activating macroautophagy/autophagy and detoxifying pathways, natural compound SFN may trigger a self-defense cellular mechanism that can effectively mitigate oxidative stress commonly associated with many metabolic and age-related diseases.

Abbreviations: ANOVA: analyzes of variance; AREs: antioxidant response elements; Baf-A1: bafilomycin A₁; BHA: butylhydroxyanisole; CAT: catechin hydrate; CCCP: carbonyl cyanide m- chlorophenylhydrazone; CLEAR: coordinated lysosomal expression and regulation; DCFH-DA: 2ʹ,7ʹ-dichlorofluorescin diacetate; FBS: fetal bovine serum; GFP: green fluorescent protein; HMOX1/HO-1: heme oxygenase 1; KD: knockdown; KEAP1: kelch like ECH associated protein 1; KO: knockout; LAMP1: lysosomal associated membrane protein 1; MCOLN1/TRPML1: mucolipin 1; ML-SA1: mucolipin-specific synthetic agonist 1; ML-SI3: mucolipin-specific synthetic inhibitor 3; MTOR: mechanistic target of rapamycin kinase; MTORC1: mechanistic target of rapamycin kinase complex 1; NAC: N-acetylcysteine; NFE2L2/NRF2: nuclear factor: erythroid 2 like 2; NPC: Niemann–Pick type C; PBS: phosphate-buffered saline; PPP2/PP2A: protein phosphatase 2; Q-PCR: real time polymerase chain reaction; ROS: reactive oxygen species; RPS6KB1/S6K1/p70S6K: ribosomal protein S6 kinase B1; SFN: sulforaphane; TFEB: transcription factor EB; WT, wild-type

氧化应激是许多病理状况的基础，包括癌症、神经变性和衰老。富含抗氧化剂的食物有助于维持细胞氧化还原稳态并减轻氧化应激，但其潜在机制尚不清楚。例如，萝卜硫素 (SFN) 是一种富含西兰花等十字花科蔬菜的亲电子化合物，是细胞抗氧化反应的有效诱导剂。NFE2L2/NRF2（核因子，类红细胞 2 样 2）是一种转录因子，通过抗氧化反应元件 (ARE) 控制多种解毒酶的表达，是 SFN 的拟议目标。NFE2L2/NRF2是 TFEB（转录因子 EB）的靶基因，TFEB 是自噬和溶酶体功能的主要调节因子，我们在此显示其可被 SFN 有效激活。SFN 通过 Ca ²⁺诱导 TFEB 核易位依赖但 MTOR（雷帕霉素激酶的机制目标）非依赖机制，通过适度增加活性氧 (ROS)。然后激活的 TFEB 会促进自噬体和溶酶体生物发生所需基因的表达，众所周知，这些基因有助于清除受损线粒体。值得注意的是，SFN 诱导的针对急性氧化剂爆发和慢性氧化应激的保护需要 TFEB 活性。因此，通过同时激活巨自噬/自噬和解毒途径，天然化合物 SFN 可以触发自卫细胞机制，可以有效减轻通常与许多代谢和年龄相关疾病相关的氧化应激。

缩写词： ANOVA：方差分析；AREs：抗氧化反应元件；Baf-A1：巴弗洛霉素 A ₁; BHA：丁基羟基茴香醚；CAT：儿茶素水合物；CCCP：羰基氰，间氯苯腙；CLEAR：协调溶酶体表达和调节；DCFH-DA：2ʹ，7ʹ-二氯荧光素二乙酸酯；FBS：胎牛血清；GFP：绿色荧光蛋白；HMOX1/HO-1：血红素加氧酶 1；KD：击倒；KEAP1：kelch like ECH相关蛋白1；KO：淘汰赛；LAMP1：溶酶体相关膜蛋白 1；MCOLN1/TRPML1：粘蛋白 1；ML-SA1：黏蛋白特异性合成激动剂 1；ML-SI3：粘蛋白特异性合成抑制剂 3；MTOR：雷帕霉素激酶的机制靶点；MTORC1：雷帕霉素激酶复合物 1 的机制靶点；NAC：N-乙酰半胱氨酸；NFE2L2/NRF2：核因子：类红细胞2像2；NPC：尼曼-匹克 C 型；PBS：磷酸盐缓冲盐水；PPP2/PP2A：蛋白磷酸酶2；Q-PCR：实时聚合酶链反应；罗斯：活性氧；RPS6KB1/S6K1/p70S6K：核糖体蛋白S6激酶B1；SFN：萝卜硫素；TFEB：转录因子EB；WT，野生型