Neuronal cell loss contributes to the pathology of acute and chronic neurodegenerative diseases, including Alzheimer’s disease (AD). It remains crucial to identify molecular mechanisms sensitizing neurons to various insults and cell death. To date, the multifunctional, autophagy-related protein Beclin 1 has been shown to be both necessary and sufficient for neuronal integrity in neurodegenerative models associated with protein aggregation. Interestingly, besides its role in cellular homeostasis, Beclin 1 has also been ascribed a role in apoptosis. This makes it critical to elucidate whether Beclin 1 regulates neuronal death and survival across neurodegenerative conditions independent of protein clearance. Here, we provide experimental evidence for a direct functional link between proteolytic cleavage of Beclin 1 and apoptotic neuronal cell loss in two independent models of neurodegeneration in vivo. Proteolytic cleavage of Beclin 1 was characterized in lysates of human AD brain samples. We developed viral tools allowing for the selective neuronal expression of the various Beclin 1 forms, including Beclin 1 cleavage products as well as a cleavage-resistant form. The effect of these Beclin 1 forms on survival and integrity of neurons was examined in models of acute and chronic neurodegeneration in vitro and in vivo. Markers of neuronal integrity, neurodegeneration and inflammation were further assessed in a Kainic acid-based mouse model of acute excitotoxic neurodegeneration and in a hAPP-transgenic mouse model of AD following perturbation of Beclin 1 in the susceptible CA1 region of the hippocampus. We find a significant increase in caspase-mediated Beclin 1 cleavage fragments in brain lysates of human AD patients and mimic this phenotype in vivo using both an excitotoxic and hAPP-transgenic mouse model of neurodegeneration. Surprisingly, overexpression of the C-terminal cleavage-fragment exacerbated neurodegeneration in two distinct models of degeneration. Local inhibition of caspase activity ameliorated neurodegeneration after excitotoxic insult and prevented Beclin 1 cleavage. Furthermore, overexpression of a cleavage-resistant form of Beclin 1 in hippocampal neurons conferred neuroprotection against excitotoxic and Amyloid beta-associated insults in vivo. Together, these findings indicate that the cleavage state of Beclin 1 determines its functional involvement in both neurodegeneration and neuroprotection. Hence, manipulating the cleavage state of Beclin 1 may represent a therapeutic strategy for preventing neuronal cell loss across multiple forms of neurodegeneration.

中文翻译：

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Beclin 1的蛋白水解裂解加剧了神经变性。

神经细胞的丢失有助于急性和慢性神经退行性疾病的病理，包括阿尔茨海默氏病（AD）。确定使神经元对各种损伤和细胞死亡敏感的分子机制仍然至关重要。迄今为止，在与蛋白质聚集相关的神经退行性模型中，多功能自噬相关蛋白Beclin 1已被证明对于神经元完整性既必要又充分。有趣的是，除了其在细胞稳态中的作用外，Beclin 1还被认为是在细胞凋亡中的作用。这使得阐明Beclin 1是否在不依赖蛋白质清除的神经变性条件下调节神经元死亡和存活变得至关重要。这里，我们提供了Beclin 1的蛋白水解切割与体内两种神经退行性变独立模型中凋亡神经元细胞丧失之间直接功能联系的实验证据。Beclin 1的蛋白水解裂解是在人类AD脑样品的裂解物中表征的。我们开发了病毒工具，可以选择性表达各种Beclin 1形式的神经元，包括Beclin 1裂解产物和抗裂解形式。在体外和体内急性和慢性神经变性模型中检查了这些Beclin 1形式对神经元存活和完整性的影响。神经元完整性的标志，在海藻酸的易感CA1区发生Beclin 1扰动后，在基于凯因酸的急性兴奋毒性神经变性的小鼠模型和AD的hAPP转基因小鼠模型中进一步评估了神经变性和炎症。我们发现人类AD患者的脑溶解产物中的半胱天冬酶介导的Beclin 1裂解片段显着增加，并使用神经毒性的兴奋性毒性和hAPP转基因小鼠模型在体内模拟该表型。出人意料的是，在两种不同的变性模型中，C末端裂解片段的过度表达加剧了神经变性。caspase活性的局部抑制改善了兴奋性中毒后神经退行性变，并阻止了Beclin 1的裂解。此外，海马神经元中抗切割形式的Beclin 1的过表达赋予神经元以对抗体内兴奋性毒性和与淀粉样蛋白相关的损伤的神经保护作用。总之，这些发现表明Beclin 1的裂解状态决定了其在神经变性和神经保护中的功能参与。因此，操纵Beclin 1的切割状态可能代表一种预防策略，可防止神经元细胞在多种形式的神经变性中丢失。