Neurosteroid Inhibition of Pyroptotic Lysis

项目摘要 上睑下垂是一种细胞死亡的程序，参与了全球主要死亡原因的发病机制。 Caspase-1家族的蛋白水解酶通过释放Gasdermin D的造孔部分启动下垂。 插入质膜导致细胞溶解。在焦油溶解过程中释放的细胞因子导致 局部和全身病理，但调节Gasdermin D孔形成和裂解的过程不是很好 明白了。我们最近确认了神经类固醇孕烯醇酮硫酸盐是一种新的焦虑症的抑制剂。 裂解，但其作用机制尚不清楚。包括孕烯醇酮硫酸盐在内的神经类固醇具有 并与质膜蛋白和脂类相互作用，这表明假设 这些分子可能会破坏Gasdermin D孔的形成。这项提案旨在了解如何 孕烯醇酮硫酸盐可防止焦油溶解，并确定其他类固醇是否具有这一活性。 我们的初步数据显示，孕烯醇酮硫酸盐可预防下垂时的松解，而不影响 炎性小体或caspase-1的上游激活。这项提案中概述的实验将测试 孕烯醇酮硫酸盐影响下睑下垂时气孔形成的假说及对Gasdermin D的评估 裂解、质膜定位和齐聚。我们还将讨论替代假设 孕烯醇酮硫酸盐可以调节细胞体积，以防止溶解或加强膜修复过程。 综上所述，这些实验的结果将为我们提供一个精确的分子理解机制。 行动，并揭示了一种新的策略，以扰乱嗜热性裂解。 类固醇的主干是由大量分子共享的，每个分子都有特定的生物学活性。我们的 初步数据表明，可能存在类固醇抑制焦链溶解的分子决定因素，作为一种 相关类固醇的效力降低。我们将测试一个合理选择的、结构截然不同的库 类固醇的能力，以防止松解症。这些实验将决定内源性的 类固醇显示出与潜在的生理相关性一致的效力。此外，这些结果将 确定抑制焦链溶解的构效关系，以促进有效的和 并定义了疾病干预的新范式。

Project Summary Pyroptosis is a program of cell death involved in the pathogenesis of leading global causes of mortality. Caspase-1 family proteases initiate pyroptosis by releasing the pore-forming portion of gasdermin D, which inserts into the plasma membrane leading to cell lysis. Cellular factors released during pyroptotic lysis cause local and systemic pathology, but processes regulating gasdermin D pore formation and lysis are not well understood. We recently identified the neurosteroid pregnenolone sulfate as a novel inhibitor of pyroptotic lysis, but its mechanism of action is not yet known. Neurosteroids including pregnenolone sulfate have a lipophilic backbone and interact with plasma membrane proteins and lipids, suggesting the hypothesis that these molecules may disrupt formation of gasdermin D pores. This proposal aims to understand how pregnenolone sulfate prevents pyroptotic lysis and determine whether other steroids share this activity. Our preliminary data demonstrate that pregnenolone sulfate prevents lysis during pyroptosis without affecting upstream activation of inflammasomes or caspase-1. The experiments outlined in this proposal will test the hypothesis that pregnenolone sulfate affects pore formation during pyroptosis and assess gasdermin D cleavage, plasma membrane localization and oligomerization. We will also address the alternative hypotheses that pregnenolone sulfate may regulate cell volume to prevent lysis or potentiate membrane repair processes. Together, the results of these experiments will inform a precise molecular understanding of the mechanism of action and reveal a novel strategy to disrupt pyroptotic lysis. The steroid backbone is shared by a large number of molecules, each with specific biological activities. Our preliminary data suggest that there may be molecular determinants for steroid inhibition of pyroptotic lysis, as a related steroid demonstrates reduced potency. We will test a rationally selected library of structurally distinct steroids for the ability to prevent pyroptotic lysis. These experiments will determine whether endogenous steroids demonstrate potency consistent with potential physiologic relevance. In addition, these results will identify the structure-activity relationship for inhibition of pyroptotic lysis to facilitate development of potent and specific molecules and define a new paradigm for disease intervention.