The phosphopeptide P140/Lupuzor, which improves the course of lupus disease in mice and patients, targets chaperone-mediated autophagy (CMA), a selective form of autophagy that is abnormally upregulated in lupus-prone MRL/lpr mice. Administered intravenously to diseased mice, P140 reduces the expression level of two major protein players of CMA, LAMP2A and HSPA8, and inhibits CMA in vitro in a cell line that stably expresses a CMA reporter. Here, we aimed to demonstrate that P140 also affects CMA in vivo and to unravel the precise cellular mechanism of how P140 interacts with the CMA process. MRL/lpr mice and CBA/J mice used as control received P140 or control peptides intravenously. Lysosome-enriched fractions of spleen or liver were prepared to examine lysosomal function. Highly purified lysosomes were further isolated and left to incubate with the CMA substrate to study at which cellular step P140 interacts with the CMA process. The data show that P140 effectively regulates CMA in vivo in MRL/lpr mice at the step of substrate lysosomal uptake and restores some alterations of defective lysosomes. For the first time, it is demonstrated that by occluding the intralysosome uptake of CMA substrates, a therapeutic molecule can attenuate excessive CMA activity in a pathological pro-inflammatory context and protect against hyperinflammation. This recovery effect of P140 on hyperactivated CMA is not only important for lupus therapy but potentially also for treating other (auto)inflammatory diseases, including neurologic and metabolic disorders, where CMA modulation would be highly beneficial.

中文翻译：

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狼疮中磷酸肽改变的自噬-溶酶体途径的体内重塑

磷酸肽P140 / Lupuzor可改善小鼠和患者的狼疮病程，其靶向分子伴侣介导的自噬（CMA），这是一种选择性形式的自噬，在易患狼疮的MRL / lpr小鼠中异常上调。对患病小鼠静脉内给药，P140降低了CMA的两个主要蛋白分子LAMP2A和HSPA8的表达水平，并在稳定表达CMA报告基因的细胞系中体外抑制了CMA。在这里，我们旨在证明P140在体内也会影响CMA，并阐明P140如何与CMA过程相互作用的确切细胞机制。用作对照的MRL / lpr小鼠和CBA / J小鼠静脉内接受P140或对照肽。准备富集脾脏或肝脏的溶酶体部分，以检查溶酶体功能。进一步分离出高度纯化的溶酶体，并与CMA底物一起孵育，以研究细胞步骤P140与CMA过程相互作用的地方。数据显示，P140在底物溶酶体摄取步骤中有效地调节了MRL / lpr小鼠体内的CMA，并恢复了有缺陷的溶酶体的某些改变。首次证明，通过阻断CMA底物的溶酶体摄取，治疗性分子可在病理性促炎环境中减弱过量的CMA活性并防止过度炎症。P140对过度活化的CMA的这种恢复作用不仅对狼疮治疗很重要，而且对治疗其他（自发性）炎症疾病（包括神经系统疾病和代谢性疾病）也很重要，在这些疾病中CMA调节将非常有益。