Lysosome integrity is essential for cell viability, and lesions in lysosome membranes are repaired by the ESCRT machinery. Here, we describe an additional mechanism for lysosome repair that is activated independently of ESCRT recruitment. Lipidomic analyses showed increases in lysosomal phosphatidylserine and cholesterol after damage. Electron microscopy demonstrated that lysosomal membrane damage is rapidly followed by the formation of contacts with the endoplasmic reticulum (ER), which depends on the ER proteins VAPA/B. The cholesterol-binding protein ORP1L was recruited to damaged lysosomes, accompanied by cholesterol accumulation by a mechanism that required VAP–ORP1L interactions. The PtdIns 4-kinase PI4K2A rapidly produced PtdIns4P on lysosomes upon damage, and knockout of PI4K2A inhibited damage-induced accumulation of ORP1L and cholesterol and led to the failure of lysosomal membrane repair. The cholesterol–PtdIns4P transporter OSBP was also recruited upon damage, and its depletion caused lysosomal accumulation of PtdIns4P and resulted in cell death. We conclude that ER contacts are activated on damaged lysosomes in parallel to ESCRTs to provide lipids for membrane repair, and that PtdIns4P generation and removal are central in this response.

中文翻译：

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通过内质网接触的胆固醇转移介导溶酶体损伤修复

溶酶体的完整性对细胞活力至关重要，溶酶体膜的损伤由 ESCRT 机制修复。在这里，我们描述了一种独立于 ESCRT 募集而激活的溶酶体修复的额外机制。脂质组学分析显示损伤后溶酶体磷脂酰丝氨酸和胆固醇增加。电子显微镜表明，溶酶体膜损伤后迅速与内质网 (ER) 形成接触，这取决于 ER 蛋白 VAPA/B。胆固醇结合蛋白 ORP1L 被募集到受损的溶酶体中，伴随着胆固醇的积累，这种机制需要 VAP-ORP1L 相互作用。PtdIns 4-激酶 PI4K2A 在受损后迅速在溶酶体上产生 PtdIns4P，PI4K2A 的敲除抑制了损伤诱导的 ORP1L 和胆固醇的积累，并导致溶酶体膜修复失败。胆固醇-PtdIns4P 转运蛋白 OSBP 也在损伤时被募集，其耗尽导致 PtdIns4P 的溶酶体积累并导致细胞死亡。我们得出结论，ER 接触在受损的溶酶体上被激活，与 ESCRT 平行，为膜修复提供脂质，而 PtdIns4P 的产生和去除是这种反应的核心。