Peroxisomes as Cellular Adaptors to Metabolic and Environmental Stress
过氧化物酶体作为代谢和环境应激的细胞适应剂

Regulation of oxidative stress by peroxisomes
过氧化物酶体对氧化应激的调节

Peroxisomes derive their name from their role in H₂O₂ metabolism. Peroxisomal respiration accounts for up to 20% of total cellular oxygen consumption and produces as much as 35% of total H₂O₂ in certain mammalian tissues [6] through the actions of multiple FAD-dependent oxidoreductases present in these organelles. To counteract the damaging effects of ROS, peroxisomes contain several antioxidant enzymes, including catalase, which reduces H₂O₂ to water. Disruption of the balance between peroxisomal ROS production and removal leads to oxidative stress and is associated with a variety of age-related human diseases, including diabetes, cancer, and neurological disorders [7–9]. For example, humans with a gain of function mutation (N237S) in Acox1 (acyl CoA oxidase 1), a peroxisomal enzyme that oxidizes VLCFA and produces H₂O₂ as a byproduct, exhibit loss of glia and neurodegeneration. Mimicking the human disease, a Drosophila model of this mutation, which promotes dimerization of the enzyme, leads to increased ROS production in glia, resulting in glial and axonal loss and increased lethality, which could be rescued by treatment with the antioxidant N-acetyl cysteine amide [10]. Increased ROS production by the active Acox1 dimer has also been implicated in DNA damage and cancer in a process dependent on sirtuin 5 (SIRT5) [11]. SIRT5 was shown to be localized in peroxisomes, where it interacts with Acox1 and promotes its desuccinylation and inhibits its activity through suppression of dimer formation. SIRT5 protein level is downregulated in hepatocellular carcinoma tumor compared to peritumor tissues, while Acox1 activity is upregulated due to increased succinylation of the β-oxidation enzyme. Knockdown of SIRT5 in human cancer cell lines increases H₂O₂ production and oxidative DNA damage in an Acox1-dependent manner [11]. It remains to be determined whether the peroxisomal localization of SIRT5, a predominantly mitochondrial and cytosolic enzyme [12], is regulated by oxidative stress.
过氧化物酶体因其在 H₂O₂ 代谢中的作用而得名。过氧化物酶体呼吸占细胞总耗氧量的 20%，并通过这些细胞器中存在的多种 FAD 依赖性氧化还原酶的作用，在某些哺乳动物组织中产生多达 35% 的总 H₂O₂ [ 6 ]。为了抵消 ROS 的破坏作用，过氧化物酶体含有多种抗氧化酶，包括过氧化氢酶，可将 H₂O₂ 还原为水。过氧化物酶体 ROS 产生和去除之间平衡的破坏会导致氧化应激，并与多种与年龄相关的人类疾病有关，包括糖尿病、癌症和神经系统疾病 [ 7 – 9 ]。例如，Acox1（酰基辅酶 A 氧化酶 1）功能获得突变（N237S）的人类，Acox1（一种氧化 VLCFA 并产生 H₂O₂ 作为副产物的过氧化物酶）表现出神经胶质细胞丧失和神经变性。模拟人类疾病，这种突变的果蝇模型促进酶的二聚化，导致神经胶质细胞中 ROS 的产生增加，导致神经胶质细胞和轴突丢失以及致死率增加，这可以通过用抗氧化剂 N-乙酰半胱氨酸酰胺处理来挽救[ 10 ]。活性 Acox1 二聚体增加 ROS 的产生也与 DNA 损伤和癌症有关，这一过程依赖于去乙醯化酶 5（SIRT5）[ 11 ]。SIRT5 被证明定位于过氧化物酶体中，它与 Acox1 相互作用并促进其去琥珀酰化并通过抑制二聚体形成来抑制其活性。 与肿瘤周围组织相比，肝细胞癌肿瘤中的 SIRT5 蛋白水平下调，而 Acox1 活性由于β氧化酶的琥珀酰化增加而上调。敲低人癌细胞系中 SIRT5 会以 Acox1 依赖性方式增加 H₂O₂ 的产生和氧化 DNA 损伤[ 11 ]。SIRT5（一种主要在线粒体和胞质 12 酶）的过氧化物酶体定位是否受氧化应激调节仍有待确定。

Genome-wide shRNA and CRISPR-Cas9 screens to discover regulators of oxidative stress identified known regulators of cellular redox balance, such as various antioxidant enzymes, but also led to the identification of several proteins involved in the peroxisomal import pathway, including Pex5, Pex13 and Pex14, as modifiers of cellular sensitivity to oxidative stress [13]. Of note, Pex5 is the import receptor of peroxisomal matrix proteins, including catalase, which contain a carboxyl terminal peroxisomal targeting sequence (PTS1) [14]. Paradoxically, inactivation of Pex5, which results in redistribution of catalase to the cytoplasm, increased resistance to oxidative stress caused by exogenous H₂O₂ treatment. Furthermore, ROS-induced phosphorylation of Pex14 at Ser232 impairs the interaction between the Pex5-Pex14 complex and catalase, confining catalase in the cytosol to counteract oxidative stress [15]. Indeed, a mutant of catalase localized to the cytoplasm was more protective against cell death caused by ROS as compared to the peroxisome-localized antioxidant enzyme [13]. This suggests that cytoplasmic catalase might be important for protection against acute oxidative stress, while peroxisomal catalase could be more involved in controlling basal oxidative stress resulting from peroxisome-generated ROS. This is consistent with previous studies suggesting that Pex5 itself is a redox-sensitive protein that exhibits reduced ability to import catalase under acute oxidative stress [16] and that the proapoptotic BCL-2 effector protein BAK promotes peroxisomal membrane permeabilization and localization of catalase in the cytosol during oxidative stress [17]. These studies support the notion that peroxisomes can function as a source or a sink of ROS, depending on physiological context [9]. The balance between ROS generation and elimination requires peroxisomal homeostasis, which in turn is maintained through a delicate balance between biogenesis and turnover of peroxisomes.
全基因组 shRNA 和 CRISPR-Cas9 筛选以发现氧化应激调节因子，鉴定了细胞氧化还原平衡的已知调节因子，例如各种抗氧化酶，但也鉴定了参与过氧化物酶体输入途径的几种蛋白质，包括 Pex5、Pex13 和 Pex14，作为细胞对氧化应激敏感性的修饰因子 [ 13 ].值得注意的是，Pex5 是过氧化物酶体基质蛋白（包括过氧化氢酶）的输入受体，过氧化氢酶含有羧基末端过氧化物酶体靶向序列（terboxyl terminal peroxisomal targeting sequence， PTS1）[ 14 ]。矛盾的是，Pex5 的失活导致过氧化氢酶重新分布到细胞质，增加了对外源性 H₂O₂ 处理引起的氧化应激的抵抗力。此外，ROS 诱导的 Pex14 在 Ser232 处的磷酸化损害了 Pex5-Pex14 复合物与过氧化氢酶之间的相互作用，将过氧化氢酶限制在细胞质中以抵消氧化应激[ 15 ]。事实上，与过氧化物酶体定位的抗氧化酶相比，定位于细胞质的过氧化氢酶突变体对 ROS 引起的细胞死亡具有更强的保护作用[ 13 ]。这表明细胞质过氧化氢酶可能对防止急性氧化应激很重要，而过氧化物酶体过氧化氢酶可能更多地参与控制过氧化物酶体产生的 ROS 引起的基础氧化应激。这与先前的研究表明，Pex5 本身是一种氧化还原敏感蛋白，在急性氧化应激下输入过氧化氢酶的能力降低[ 16 ]，并且促凋亡 BCL-2 效应蛋白 BAK 在氧化应激期间促进过氧化物酶体膜透化和过氧化氢酶在细胞质中的定位[ 17 ]。 这些研究支持过氧化物酶体可以作为 ROS 的来源或汇的观点，具体取决于生理背景[ 9 ]。ROS 生成和消除之间的平衡需要过氧化物酶体稳态，而过氧化物酶体稳态又通过过氧化物酶体的生物发生和周转之间的微妙平衡来维持。

Turnover of old or dysfunctional peroxisomes has also been linked to increased peroxisomal ROS production. In this regard, ATM (Ataxia-telangiectasia mutated) kinase, a redox sensitive regulator of nuclear DNA damage response [18, 19], was recently shown to interact with Pex5 and translocate to peroxisomes to regulate pexophagy in response to ROS [20]. Activated ATM phosphorylates Pex5 at Ser 141, which in turn promotes Pex5 ubiquitination at Lys 209. Peroxisomes containing ubiquitinated Pex5 are recognized by the autophagy adaptor p62 and targeted for degradation [20]. It remains unclear whether ATM is activated by ROS endogenously produced by peroxisomes. However, supporting the possibility that endogenous ROS promotes autophagic degradation of peroxisomes, activation of pexophagy caused by knockout of the heat shock protein HSPA9 has been linked to elevated accumulation of peroxisomal ROS detected using peroxisome-localized HyPer, a redox-sensitive fluorescent protein that can detect H₂O₂ [21]. One potential beneficial effect of ROS-induced pexophagy mediated by ATM could be the restoration of cellular redox balance to prevent ROS-induced DNA damage. In line with this notion, knockdown of catalase leads to the impairment of peroxisomal ROS removal and causes ROS-induced pexophagy, which can be rescued by the addition of the antioxidant agent N-acetyl-L-cysteine [22].
旧的或功能失调的过氧化物酶体的周转也与过氧化物酶体 ROS 产生增加有关。在这方面，ATM（共济失调-毛细血管扩张突变）激酶是核 DNA 损伤反应的氧化还原敏感调节因子 [ 18 ， 19 ]，最近被证明与 Pex5 相互作用并易位到过氧化物酶体以调节响应 ROS 的 pexophagagy [ 20 ]。激活的 ATM 在 Ser 141 处磷酸化 Pex5，进而促进 Lys 209 处的 Pex5 泛素化。含有泛素化 Pex5 的过氧化物酶体被自噬接头 p62 识别并靶向降解[ 20 ]。目前尚不清楚 ATM 是否由过氧化物酶体内源性产生的 ROS 激活。然而，支持内源性 ROS 促进过氧化物酶体自噬降解的可能性，由热休克蛋白 HSPA9 敲除引起的 pexophagage 激活与使用过氧化物酶体定位的 HyPer 检测到的过氧化物酶体 ROS 积累升高有关，HyPer 是一种氧化还原敏感荧光蛋白，可以检测 H₂O₂ [ 21 ].ATM 介导的 ROS 诱导的 pexophagy 的一个潜在有益作用可能是恢复细胞氧化还原平衡，以防止 ROS 诱导的 DNA 损伤。根据这一观点，过氧化氢酶的敲低会导致过氧化物酶体 ROS 去除受损，并引起 ROS 诱导的 pexophagage，这可以通过添加抗氧化剂 N-乙酰基-L-半胱氨酸来挽救[ 22 ]。

Role of peroxisomes in metabolic adaptation to hypoxic stress
过氧化物酶体在代谢适应缺氧应激中的作用

Oxygen is a critical substrate in cellular metabolism and bioenergetics. Inadequate availability of oxygen to tissues, or hypoxia, is experienced in various physiological and pathological states. For instance, tissue hypoxia is a prominent feature of most solid tumors that promotes tumor aggressiveness and metastasis [23, 24]. A genome-wide CRISPR-Cas9 growth screen by Jain et al. [25] to discover regulators of cell fitness in high and low oxygen conditions led to the identification of an essential role for peroxisomes in metabolic adaptation to hypoxic stress. Notably, the authors observed that peroxisomal biogenesis genes Pex2, Pex5, Pex7, Pex10, Pex12, and Pex13 and plasmalogen synthesis genes GNPAT, AGPS, FAR1/2 and TMEM189 were selectively essential for growth in 1% O₂ versus 21% O₂. Plasmalogens are peroxisome-derived phospholipids which have a vinyl ether bond at the sn-1 position as opposed to an ester bond in the conventional phospholipids (Box 3). The cell growth defect under hypoxic stress was rescued with the addition of the unsaturated fattyacid oleate, but worsened with the saturated fatty acid palmitate [25]. This is notable since previous studies suggest that stearoyl-CoA desaturase (SCD), an enzyme that introduces a single double bond in the Δ9 position of saturated fatty acids, is an oxygen-sensitive protein inhibited by hypoxia. The resulting increased ratio of saturated to unsaturated fatty acids leads to impaired cell growth, presumably due to increased membrane rigidity [26, 27]. This led Jain et al. to propose that plasmalogens, which have been implicated in membrane fluidity [28], are beneficial in hypoxia to reduce membrane rigidity. Their lipidomics analysis indicated that biosynthesis of these peroxisome-derived lipids is increased in hypoxia [25]. It remains to be determined whether plasmalogens directly influence cellular fitness in the context of hypoxic stress via control of membrane fluidity. It is possible that other mechanisms might also be at play. For example, plasmalogens are thought to function as endogenous antioxidants [29, 30], which could affect hypoxia-related lipid peroxidation [31]. In line with this possibility, a genome-wide CRISPR-Cas9 screen coupled with lipidomic profiling to discover factors that impact susceptibility to ferroptosis led to the identification of polyunsaturated ether lipids, including plasmalogens, as substrates for lipid peroxidation that promote induction of ferroptosis, which carcinoma cells can evade through downregulation of these lipids [32].
氧气是细胞代谢和生物能量学中的关键底物。在各种生理和病理状态下，组织氧气供应不足或缺氧。例如，组织缺氧是大多数实体瘤的一个突出特征，可促进肿瘤的侵袭性和转移 [ 23 ， 24 ]。Jain 等人 [ 25 ] 通过全基因组 CRISPR-Cas9 生长筛选，以发现高氧和低氧条件下细胞适应性的调节因子，从而确定了过氧化物酶体在代谢适应缺氧应激中的重要作用。值得注意的是，作者观察到过氧化物酶体生物发生基因 Pex2、Pex5、Pex7、Pex10、Pex12 和 Pex13 以及缩醛磷脂合成基因 GNPAT、AGPS、FAR1/2 和 TMEM189 对于 1% O₂ 与 21% O₂ 的生长具有选择性必要性。疟原蛋白是过氧化物酶体衍生的磷脂，在 sn-1 位具有乙烯基醚键，而不是传统磷脂中的酯键 （ Box 3 ）。添加不饱和脂肪酸油酸酯可挽救缺氧胁迫下的细胞生长缺陷，但加入饱和脂肪酸棕榈酸酯可加重[ 25 ]。这一点值得注意，因为先前的研究表明，硬脂酰辅酶 A 去饱和酶 （SCD） 是一种在饱和脂肪酸的 Δ9 位引入单双键的酶，是一种因缺氧而抑制的氧敏感蛋白质。由此产生的饱和脂肪酸与不饱和脂肪酸比例增加导致细胞生长受损，可能是由于膜刚度增加 [ 26 ， 27 ]。这导致 Jain 等人提出，与膜流动性有关的缩醛磷脂[ 28 ]有利于缺氧，从而降低膜刚度。 他们的脂质组学分析表明，这些过氧化物酶体衍生脂质的生物合成在缺氧时增加 [ 25 ]。在缺氧应激背景下，疟原是否通过控制膜流动性直接影响细胞适应性仍有待确定。其他机制也可能在起作用。例如，缩醛磷脂被认为具有内源性抗氧化剂的作用 [ 29 ， 30 ]，这可能会影响缺氧相关的脂质过氧化 [ 31 ]。与这种可能性一致，全基因组 CRISPR-Cas9 筛选与脂质组学分析相结合，以发现影响铁死亡易感性的因素，从而鉴定出多不饱和醚脂质，包括缩醛磷脂，作为脂质过氧化的底物，促进诱导铁死亡，癌细胞可以通过下调这些脂质来逃避铁死亡[ 32 ]。

It is remarkable that while acute exposure of cells to low oxygen conditions does not decrease peroxisome abundance [25], sustained activation of hypoxia via genetic inactivation of a critical regulatory factor dramatically decreases the number of peroxisomes [33]. Hypoxia is regulated by the hypoxia inducible factor (HIF) family of transcription factors, which are active under hypoxia but targeted for degradation by the Von Hippel-Lindau (VHL) protein under oxygen replete conditions. Activated HIF transcriptionally reprogram cells to adapt to the low oxygen tension environment [34]. Interestingly, the stabilization of HIF2α resulting from VHL knockout promotes pexophagy, leading to a dramatic reduction of peroxisome abundance [33]. The precise reason for the conflicting effects on peroxisome abundance remains unclear. However, chronic hypoxia has been shown to protect against the effects of acute hypoxia [35, 36]. Thus, it is possible that induction of pexophagy as a result of sustained hypoxic stress might reflect a feedback mechanism to maintain peroxisome homeostasis.
值得注意的是，虽然细胞急性暴露于低氧条件下不会降低过氧化物酶体丰度[ 25 ]，但通过关键调节因子的遗传失活来持续激活缺氧会显着减少过氧化物酶体的数量[ 33 ]。缺氧受缺氧诱导因子 （HIF） 家族转录因子的调节，这些转录因子在缺氧条件下具有活性，但在富氧条件下被 Von Hippel-Lindau （VHL） 蛋白降解。激活的 HIF 转录重编程细胞以适应低氧张压环境[ 34 ]。有趣的是，VHL 敲除导致的 HIF2α的稳定促进了 pexophagage，导致过氧化物酶体丰度急剧降低[ 33 ]。对过氧化物酶体丰度相互影响的确切原因尚不清楚。然而，慢性缺氧已被证明可以防止急性缺氧的影响 [ 35 ， 36 ]。因此，由于持续的缺氧应激而诱导 pexophagy 可能反映了维持过氧化物酶体稳态的反馈机制。

Peroxisomal regulation of lipid homeostasis during nutrient stress
营养胁迫期间脂质稳态的过氧化物酶体调节

Adipose tissue stores excess energy as triglycerides within lipid droplets. This pool of intracellular lipid stores can be mobilized during nutrient deprivation through a process called lipolysis, which involves induction of β-adrenergic receptor signaling, resulting in the activation of a cAMP and protein kinase A (PKA)-dependent pathway [37]. Lipolysis is mediated by a series of lipases, including adipose triglyceride lipase (ATGL), an enzyme that catalyzes the first and rate-limiting step in the process, hydrolyzing triglycerides to diglycerides. Activities of lipolytic enzymes are regulated by nutritional status [37], but the mechanism through which these enzymes translocate onto lipid droplet and catalyze lipolysis remains ill-defined.
脂肪组织将多余的能量作为甘油三酯储存在脂滴中。在营养剥夺期间，这种细胞内脂质储存可以通过称为脂肪分解的过程进行调动，该过程涉及诱导β肾上腺素能受体信号传导，从而激活 cAMP 和蛋白激酶 A（PKA）依赖性途径[ 37 ]。脂肪分解由一系列脂肪酶介导，包括脂肪甘油三酯脂肪酶 （ATGL），这是一种催化过程中的第一步和限速步骤的酶，将甘油三酯水解为甘油二酯。脂肪分解酶的活性受营养状况的调节[ 37 ]，但这些酶易位到脂滴上并催化脂肪分解的机制仍然不明确。

A recent study by Kong et al. [38] identified a role for peroxisomes in fasting-induced lipolysis in adipose tissue.Peroxisomes have long been known to associate with lipid droplets [39], but the physiological significance of this interaction has remained unclear. Kong et al. discovered that fasting stimulates physical interaction between peroxisomes and lipid droplets and induces fatty acid liberation via lipolysis in white adipose tissue (WAT) [38]. Similar results were observed in 3T3-L1 adipocytes treated with isoproterenol, a β-adrenergic receptor agonist, to mimic the fasting response. Mechanistically, the authors demonstrated that fasting promotes interaction between peroxisomes and lipid droplets by increasing kinesin-like protein KIFC3-dependent movement of peroxisomes toward lipid droplets. Moreover, they reported that the peroxisomal biogenesis factor Pex5 interacts with ATGL and recruits it to peroxisome-lipid droplet contact sites in a manner dependent on PKA activation [38]. However, it is unclear how Pex5 interacts with ATGL since the lipolytic enzyme lacks a PTS1. It is also unclear what the fate is of the fatty acids liberated by peroxisome-mediated lipolysis.
Kong 等[ 38 ]最近的一项研究确定了过氧化物酶体在空腹诱导的脂肪组织脂肪分解中的作用。长期以来，人们一直知道过氧化物酶体与脂滴结合[ 39 ]，但这种相互作用的生理意义仍不清楚。Kong 等发现，禁食会刺激过氧化物酶体和脂滴之间的物理相互作用，并通过脂肪分解在白色脂肪组织（white adiposition organization， WAT）中诱导脂肪酸释放[ 38 ]。在用异丙肾上腺素（一种β-肾上腺素能受体激动剂）处理的 3T3-L1 脂肪细胞中观察到类似的结果，以模拟禁食反应。从机制上讲，作者证明，禁食通过增加过氧化物酶体向脂滴的驱动蛋白样蛋白 KIFC3 依赖性运动来促进过氧化物酶体和脂滴之间的相互作用。此外，他们报告说，过氧化物酶体生物发生因子 Pex5 与 ATGL 相互作用，并以依赖于 PKA 激活的方式将其募集到过氧化物酶体-脂滴接触位点 [ 38 ]。然而，尚不清楚 Pex5 如何与 ATGL 相互作用，因为脂肪分解酶缺乏 PTS1。目前还不清楚过氧化物酶体介导的脂肪分解释放的脂肪酸的命运是什么。

Besides lipolysis, fatty acids could be liberated from triglycerides by lipophagy, a subtype of autophagy that involves encapsulation of lipid droplets within a double-membrane autophagosome, which fuses with a lysosome, followed by hydrolysis of triglycerides by lysosomal acid lipase A (LAL) [40]. Starvation activates the autophagy pathway, but paradoxically autophagic degradation of lipid droplets is normally not sufficient to prevent hepatic steatosis that results from prolonged nutrient deprivation. Acetyl-CoA is a key metabolic regulator of autophagy whose depletion is sufficient to promote autophagy activation, albeit through a previously undefined mechanism [41].
除脂肪分解外，脂肪酸还可以通过自噬从甘油三酯中释放出来， 脂肪自噬是自噬的一种亚型，涉及将脂滴包裹在双膜自噬体内，与溶酶体融合，然后通过溶酶体酸性脂肪酶 A（LAL） 40 水解甘油三酯[ ]。饥饿会激活自噬途径，但矛盾的是，脂滴的自噬降解通常不足以防止因长期营养剥夺而导致的肝脂肪变性。乙酰辅酶 A 是自噬的关键代谢调节剂，其消耗足以促进自噬激活，尽管是通过先前未定义的机制[ 41 ]。

Recent studies suggest that peroxisomal β-oxidation is a major source of cytosolic acetyl CoA that limits lipophagy activation in the liver under nutrient deprivation [42]. Hepatocyte-specific genetic inactivation of the peroxisomal β-oxidation enzyme Acox1 prevents against starvation-induced fatty liver in mice. Mechanistically, acetyl-CoA derived from Acox1-mediated fatty acid oxidation promotes acetylation of Raptor [42], a component of the mTOR (mechanistic target of rapamycin kinase) complex 1 (mTORC1) [43]. This growth and metabolism regulatory complex senses nutrients and inhibits autophagy under nutrient sufficiency by phosphorylating various autophagy-related proteins, including ULK1 (Unc-51-like autophagy-activating kinase 1) [44], which promotes autophagy initiation and autophagosome formation. Raptor regulates subcellular localization of mTOR and promotes recruitment of its substrates [45]. As a consequence of hepatic Acox1 deficiency, lysosomal localization and activation of mTOR is impaired, resulting in decreased autophagy-inhibitory Ser757 phosphorylation of ULK1 and subsequent activation of lipophagy and protection against fatty liver [42]. The activation of mTORC1 was recently shown to also be impaired in peroxisome-deficient Chinese hamster ovary cells [46]. With regard to peroxisomal β-oxidation-mediated inhibition of mTORC1, these studies suggest a peroxisome-lysosome metabolic link restricts autophagic degradation of lipid droplets in the fasted state and controls hepatic lipid homeostasis. It is noteworthy that this is in contrast to the fasting-induced peroxisome-lipid droplet interaction discussed above, which enhances fatty acid liberation via the promotion of lipolysis in adipose tissue [38] (Figure 2). These opposing effects of peroxisomes on lipid hydrolysis suggest that peroxisomal processes are calibrated in a tissue-specific manner based on nutritional status. For example, fasting increases Acox1 gene expression in the liver but not in WAT [47].
最近的研究表明，过氧化物酶体β氧化是胞质乙酰辅酶 A 的主要来源，在营养剥夺下限制肝脏的噬脂激活[ 42 ]。过氧化物酶体β氧化酶 Acox1 的肝细胞特异性遗传失活可预防饥饿诱导的小鼠脂肪肝。从机制上讲，源自 Acox1 介导的脂肪酸氧化的乙酰辅酶 A 促进 Raptor[ 42 ]的乙酰化，Raptor 是 mTOR（雷帕霉素激酶机制靶标）复合物 1（mTORC1）的组分[ 43 ]。这种生长和代谢调节复合物通过磷酸化各种自噬相关蛋白（包括 ULK1（Unc-51 样自噬激活激酶 1）[ 44 ]来感知营养物质并在营养充足的情况下抑制自噬，从而促进自噬启动和自噬体形成。Raptor 调节 mTOR 的亚细胞定位并促进其底物的募集 [ 45 ]。由于肝脏 Acox1 缺乏，溶酶体定位和 mTOR 的激活受损，导致 ULK1 的自噬抑制性 Ser757 磷酸化减少，随后激活噬脂作用并预防脂肪肝 [ 42 ]。最近显示，过氧化物酶体缺陷的中国仓鼠卵巢细胞中 mTORC1 的激活也受损[ 46 ]。关于过氧化物酶体β氧化介导的 mTORC1 抑制，这些研究表明过氧化物酶体-溶酶体代谢联系限制了禁食状态下脂滴的自噬降解并控制肝脂稳态。 值得注意的是，这与上面讨论的空腹诱导的过氧化物酶体-脂滴相互作用形成鲜明对比，后者通过促进脂肪组织中的脂肪分解来增强脂肪酸游离 [ 38 ] （ Figure 2 ）。过氧化物酶体对脂质水解的这些相反作用表明，过氧化物酶体过程是根据营养状况以组织特异性方式校准的。例如，禁食会增加肝脏中 Acox1 基因的表达，但不会增加 WAT 中的 Acox1 基因表达[ 47 ]。

Peroxisome-derived lipids and the regulation of thermogenesis in response to cold exposure
过氧化物酶体衍生的脂质和响应冷暴露的产热调节

One of the challenges for euthermic animals is to maintain their body temperature in a cold environment. Cold-induced heat production in mammals occurs through skeletal muscle shivering [48] or by non-shivering thermogenesis, which is mediated primarily by brown adipose tissue (BAT) and the related beige adipocytes that appear within subcutaneous WAT in response to prolonged cold exposure or β-adrenergic stimulation [49]. Brown and beige fat cells are enriched in mitochondria and specialize in oxidizing fatty acids and glucose to generate heat via mitochondrial uncoupling. In addition to promoting thermogenesis through uncoupled respiration, mitochondria are highly dynamic organelles that regulate the thermogenic capacity of adipocytes through their ability to undergo fission in response to β-adrenergic stimulation. Fragmented and circular mitochondria are thought to exhibit increased uncoupling of oxidative phosphorylation by directing nutrient oxidation toward heat production instead of ATP synthesis [50].
真热动物面临的挑战之一是在寒冷的环境中保持体温。哺乳动物的冷诱发热量产生是通过骨骼肌颤抖[ 48 ]或非颤抖产热发生的，这主要由棕色脂肪组织（brown adiposition organization， BAT）和皮下 WAT 内出现的相关米色脂肪细胞介导，以响应长时间的寒冷暴露或β肾上腺素能刺激 49 [ ]。棕色和米色脂肪细胞富含线粒体，专门氧化脂肪酸和葡萄糖，通过线粒体解偶联产生热量。除了通过非偶联呼吸促进产热外，线粒体还是高度动态的细胞器，通过脂肪细胞响应β肾上腺素能刺激而发生裂变的能力来调节脂肪细胞的产热能力。碎片化和环状线粒体被认为通过将营养物质氧化引导至热量产生而不是 ATP 合成，表现出氧化磷酸化解偶联的增加 [ 50 ]。

Peroxisomes are also a dynamic organelle whose abundance in brown adipocytes increases with cold exposure [51, 52]. Recent studies indicate that cold-induced peroxisomal biogenesis is mediated by the thermogenic transcriptional co-regulator PRDM16 [53], suggesting that the transcriptional regulation of thermogenesis and de novo peroxisome formation is linked. Consistent with a role for peroxisomes in thermogenesis, disruption of peroxisomal biogenesis through adipose-specific knockout of the critical peroxisomal biogenesis factor Pex16 results in cold intolerance in mice due to inhibition of cold-induced mitochondrial fission. Mechanistically, the loss of peroxisomes decreases the mitochondrial membrane content of plasmalogens. Treatment of mice with alkyl glycerol, a plasmalogen precursor, restores cold-induced mitochondrial fragmentation and thermogenesis in Pex16 knockout mice, suggesting that peroxisomes channel lipids to mitochondria in brown and beige adipocytes to regulate mitochondrial dynamics and thermogenesis [53]. Future research will be required to understand the precise mechanism through which plasmalogens affect mitochondrial fission. One potential explanation is that these peroxisome-derived lipids are required to maintain mitochondrial membrane fluidity necessary for cold-induced mitochondrial fission. It is also possible that plasmalogens assist in the recruitment of factors involved in mitochondrial dynamics.
过氧化物酶体也是一种动态细胞器，其棕色脂肪细胞中的丰度随着寒冷暴露而增加 [ 51 ， 52 ]。最近的研究表明，冷诱导的过氧化物酶体生物发生是由产热转录共调节因子 PRDM16 介导的[ 53 ]，这表明产热的转录调节与从头过氧化物酶体形成有关。与过氧化物酶体在产热中的作用一致，通过脂肪特异性敲除关键的过氧化物酶体生物发生因子 Pex16 破坏过氧化物酶体生物发生，由于抑制冷诱导的线粒体裂变，导致小鼠不耐寒。从机制上讲，过氧化物酶体的丢失会降低缩醛磷脂的线粒体膜含量。用烷基甘油（一种缩醛磷前体）处理小鼠可恢复 Pex16 基因敲除小鼠中冷诱导的线粒体片段化和产热，这表明过氧化物酶体将脂质引导到棕色和米色脂肪细胞中的线粒体以调节线粒体动力学和产热 [ 53 ]。未来的研究将需要了解等离子体影响线粒体裂变的精确机制。一种可能的解释是，这些过氧化物酶体衍生的脂质是维持冷诱导线粒体裂变所必需的线粒体膜流动性所必需的。缩醛磷脂也有可能有助于招募参与线粒体动力学的因子。

Peroxisomal regulation of noise-induced hearing loss
过氧化物酶体调节噪声性听力损失

In the modern industrial society, noise is another major environmental stressor. Overexposure to noise promotes ROS production and oxidative damage, leading to auditory hair cell damage and hearing loss [54, 55], while antioxidant therapy has been shown to prevent hearing loss in mice [56]. Consistent with the important role of peroxisomes in ROS metabolism, polymorphisms in the peroxisomal antioxidant enzyme catalase are associated with noise-induced hearing loss (NIHL) in humans [57]. The initial evidence for a direct role of peroxisomes in NIHL was provided by the discovery that Pejvakin (also called DFNB59), a poorly characterized protein genetically linked to sensorineural hearing loss, is associated with peroxisomes, particularly with peroxisomal protrusions and “string-of-beads” structures indicative of peroxisome growth and fission [55]. Like humans with mutations in the gene encoding Pejvakin (PVJK), Pejvakin-deficient mice are hypervulnerable to noise. Embryonic fibroblasts from Pjvk^+/+ mice stressed with H₂O₂ show a significant increase in peroxisome number, but those from Pjvk^−/− mice remain unchanged. Furthermore, Pjvk^−/− mice only display structural peroxisome abnormalities in hair cells after hearing loss onset due to the oxidative stress caused by noise exposure [55].
在现代工业社会中，噪音是另一个主要的环境压力源。过度暴露于噪音会促进 ROS 的产生和氧化损伤，导致听觉毛细胞损伤和听力损失 [ 54 ， 55 ]，而抗氧化疗法已被证明可以预防小鼠听力损失 [ 56 ]。与过氧化物酶体在 ROS 代谢中的重要作用一致，过氧化物酶体抗氧化酶过氧化氢酶的多态性与人类噪声性听力损失（noise-induced hearing loss， NIHL）有关[ 57 ]。过氧化物酶体在 NIHL 中直接作用的初步证据是发现 Pejvakin（也称为 DFNB59）是一种与感音神经性听力损失有遗传联系的特征不佳的蛋白质，与过氧化物酶体有关，特别是与过氧化物酶体突起和指示过氧化物酶体生长和裂变的“串珠”结构有关 [ 55 ].与编码 Pejvakin （PVJK） 基因突变的人类一样，Pejvakin 缺陷小鼠对噪音非常敏感。用 H₂O₂ 胁迫的 Pjvk ^{+ / +} 小鼠的胚胎成纤维细胞显示出过氧化物酶体数量的显着增加，但来自 Pjvk^−/- 小鼠的胚胎成纤维细胞保持不变。此外，由于噪声暴露引起的氧化应激，Pjvk^−/− 小鼠在听力损失发作后仅在毛细胞中表现出结构性过氧化物酶体异常[ 55 ]。

Additional mechanistic studies suggest that following sound-induced oxidative stress, auditory cells undergo autophagic degradation of damaged peroxisomes through pexophagy prior to peroxisome proliferation. Pejvakin contains a functional LC3-interacting region (LIR) and directly recruits the autophagosomal marker LC3B to peroxisomes in response to H₂O₂ treatment in HepG2 cells to facilitate pexophagy and pave the way for peroxisome proliferation [58]. Taken together, these studies establish a key pathway in which pejvakin-mediated peroxisome degradation and proliferation protect auditory hair cells against noise-induced oxidative stress and reveal that the DFNB59 form of deafness is a pexophagy disorder. As discussed above, activation of pexophagy involves Pex5-dependent peroxisomal recruitment of the ATM kinase [20]. It is unclear whether inactivation of PVJK disrupts the Pex5/ATM axis. Nevertheless, this work opens up new therapeutic avenues as antioxidant supplementation and AAV-mediated gene therapy appear to be effective treatments for the disease [55].
其他机制研究表明，在声音诱导的氧化应激之后，听觉细胞在过氧化物酶体增殖之前通过异食对受损的过氧化物酶体进行自噬降解。Pejvakin 含有功能性 LC3 相互作用区（LIR），并直接将自噬体标志物 LC3B 募集到过氧化物酶体中，以响应 HepG2 细胞中的 H₂O₂ 处理，以促进 pexophagage 并为过氧化物酶体增殖铺平道路[ 58 ]。总之，这些研究建立了一条关键途径，其中 pejvakin 介导的过氧化物酶体降解和增殖保护听觉毛细胞免受噪音诱导的氧化应激，并揭示 DFNB59 形式的耳聋是一种吞噬障碍。如上所述，pexophagage 的激活涉及 ATM 激酶的 Pex5 依赖性过氧化物酶体募集 [ 20 ]。目前尚不清楚 PVJK 的失活是否会破坏 Pex5/ATM 轴。然而，这项工作开辟了新的治疗途径，因为抗氧化剂补充剂和 AAV 介导的基因疗法似乎是治疗该疾病的有效方法 [ 55 ]。

Role of peroxisomes in innate immune response to viral and bacterial infections
过氧化物酶体在病毒和细菌感染的先天免疫反应中的作用

Besides extreme physical environment, pathogens, such as viruses and bacteria also contribute to environmental stress in animals. Innate immune system is the first line of defense against infections. For viral infections, the innate immune response involves recognition of viral nucleic acids by pathogen recognition receptors, including cytosolic receptors such as RIG-I-like receptors (RLRs) that activate mitochondrial antiviral signaling protein (MAVS) to destroy viruses [59]. Although originally identified as a mitochondrial membrane protein that activates NF-kB and interferon regulatory factors, resulting in the production of pro-inflammatory cytokines and type I interferons (IFN), MAVS is also located at the peroxisomal membrane and triggers expression of type III IFN and IFN-stimulated genes upon viral infections [60]. The products of IFN-stimulated genes have antiviral effects, which destroy the virus or inhibit viral replication [61]. However, some viruses have evolved a strategy to escape the antiviral effect mediated by peroxisomes. For instance, the flavivirus capsid protein promotes sequestration and degradation of Pex19, a cytoplasmic chaperone of peroxisomal membrane proteins, leading to reduction of peroxisome number and impairment of early antiviral signaling [62]. Recent studies suggest that enveloped viruses, such as human cytomegalovirus (HCMV) and herpes simplex virus type 1 (HSV-1) do not inhibit peroxisomal biogenesis, but rather increase it to support their replication [63]. The authors suggest that this leads to increased production of plasmalogens necessary for secondary envelopment and the assembly of infectious particles. However, since peroxisomal lipid synthesis also contributes to the production of conventional diacyl phospholipids [64], additional work is required to determine if plasmalogens are directly necessary for envelopment.
除了极端的物理环境外，病毒和细菌等病原体也会导致动物的环境压力。先天免疫系统是抵御感染的第一道防线。对于病毒感染，先天免疫应答涉及病原体识别受体对病毒核酸的识别，包括细胞质受体，如 RIG-I 样受体（RLR），可激活线粒体抗病毒信号蛋白（mitochondrial antiviral signaling protein， MAVS）以破坏病毒[ 59 ]。虽然 MAVS 最初被鉴定为一种线粒体膜蛋白，可激活 NF-kB 和干扰素调节因子，从而产生促炎细胞因子和 I 型干扰素（IFN），但 MAVS 也位于过氧化物酶体膜上，并在病毒感染时触发 III 型干扰素和 IFN 刺激基因的表达[ 60 ]。IFN 刺激基因的产物具有抗病毒作用，可破坏病毒或抑制病毒复制[ 61 ]。然而，一些病毒已经进化出一种策略来逃避过氧化物酶体介导的抗病毒作用。例如，黄病毒衣壳蛋白促进过氧化物酶体膜蛋白的细胞质伴侣 Pex19 的螯合和降解，导致过氧化物酶体数量减少和早期抗病毒信号传导受损[ 62 ]。最近的研究表明，包膜病毒，如人类巨细胞病毒（human cytomegalovirus， HCMV）和单纯疱疹病毒 1 型（herpes simplex virus type 1， HSV-1）不会抑制过氧化物酶体生物发生，而是会增加过氧化物酶体的生物发生以支持其复制[ 63 ]。作者认为，这导致二次包膜和传染性颗粒组装所必需的缩醛磷脂的产生增加。 然而，由于过氧化物酶体脂质的合成也有助于产生常规的二酰磷脂[ 64 ]，因此需要额外的工作来确定缩醛磷脂是否直接需要包膜。

The peroxisomal role in innate immune response is not limited to only viral infections, but extends to bacterial infections as well. Microbial invaders are destroyed through phagocytosis, in which specialized cells use plasma membrane to engulf pathogens, giving rise to an internal compartment called the phagosome. The phagosome fuses with lysosome, forming a phagolysosome, which degrades pathogens [65]. A study by Di Cara et al. has identified a role for peroxisomes in phagocytosis [66]. Peroxisomes are localized to the sites of phagosome formation in Drosophila S2 cells and knockdown of Pex5 or Pex7 results in reduced bacterial uptake via phagocytosis. Peroxisome deficiency leads to disorganization of the actin network and a defect in phagocytic cup formation, coupled with impaired ROS metabolism. Overexpression of catalase in peroxisome deficient cells or treatment with phosphatidylinositols and DHA, lipids thought to be produced in peroxisomes, rescues phagocytosis [66]. These studies suggest that peroxisomal function in ROS and lipid metabolism is important for phagocytosis, but the detailed molecular mechanism remains to be discovered. Peroxisomal lipid metabolism has also been implicated in bactericidal activity through a different mechanism involving FAMIN, a laccase domain-containing protein that partially localizes to the cytosolic surface of peroxisomes through its interaction with fatty acid synthase. FAMIN regulates β-oxidation of de novo synthesized fatty acids. FAMIN-deficient macrophages have impaired mitochondrial ROS production, a defect in inflammasome activation and reduced bacterial clearance [67].
过氧化物酶体在先天免疫反应中的作用不仅限于病毒感染，还延伸到细菌感染。微生物入侵者通过吞噬作用被消灭，其中特化细胞利用质膜吞噬病原体，产生一个称为吞噬体的内部隔室。吞噬体与溶酶体融合，形成吞噬溶酶体，降解病原体 [ 65 ]。Di Cara 等人的一项研究已经确定了过氧化物酶体在吞噬作用中的作用[ 66 ]。过氧化物酶体定位于果蝇 S2 细胞中吞噬体形成的位点，敲低 Pex5 或 Pex7 导致通过吞噬作用减少细菌摄取。过氧化物酶体缺乏会导致肌动蛋白网络紊乱和吞噬杯形成缺陷，以及 ROS 代谢受损。过氧化物酶体缺陷细胞中过氧化氢酶的过度表达或用磷脂酰肌醇和 DHA（被认为是在过氧化物酶体中产生的脂质）处理，可以挽救吞噬作用 [ 66 ]。这些研究表明，ROS 和脂质代谢中的过氧化物酶体功能对吞噬作用很重要，但详细的分子机制仍有待发现。过氧化物酶体脂质代谢也通过涉及 FAMIN 的不同机制与杀菌活性有关，FAMIN 是一种含有漆酶结构域的蛋白质，通过与脂肪酸合酶的相互作用部分定位到过氧化物酶体的胞质表面。FAMIN 调节从头合成脂肪酸的β氧化。FAMIN 缺陷巨噬细胞会损害线粒体 ROS 的产生，这是炎症小体激活的缺陷和细菌清除率降低[ 67 ]。