To the edge of necroptosis and back

摘要 坏死性下垂是细胞死亡的一种形式，由FADD-caspase-8-flip复合体抑制，其中 两种蛋白激酶RIPK1和RIPK3相互作用，促进或抑制细胞的坏死性死亡。激活的RIPK3 磷酸化假性激酶MLKL，它通过在质膜上的作用而介导坏死性下垂。 我们已经探索了围绕MLKL激活的晚期事件，并发现在血浆丢失之前 膜完整，细胞死亡是可逆的。我们已经发现了ESCRT III途径，它在 胞质转运、胞质分裂和胞吐作用，以对抗活性MLKL的膜效应。 这导致受损质膜的“气泡”从细胞表面释放出来，维持细胞的生存。 生死存亡。我们的中心假设，也就是这个应用的基础，是坏死性下垂通过 MLKL的激活及其在质膜上的作用，以及这一后期过程的调节允许其他 事件，如由坏死性刺激诱导的蛋白质表达，从而影响到 死亡细胞对身体的影响。基于这一假设，我们将问：1.中国的后期事件是如何 坏死下垂调节细胞死亡和存活？在这里，我们将确定ESCRT III的组成部分 负责MLKL激活后气泡的形成，以及它们在激活MLKL时的“复苏”中的作用 随后被扰乱了。我们将确定ESCRT III如何发挥作用来抑制坏死性下垂 没有caspase抑制的诱导剂，并探索RIPK3活性是否独立于MLKL调节 ESCRT III途径。2.是否存在MLKL激活和坏死性下垂的非典型途径？而当 几种坏死性下垂的诱因已经确定，我们认为还有其他较少被研究的诱因。 我们已经确定DNA损伤是坏死性下垂的诱因，并将探索这种情况是如何发生的。此外，我们 已经发现了独立于RIPK3激活MLKL和坏死性下垂的药物，并将确定 这种非正则激活是如何发生的。3.坏死性下垂的晚期事件的后果是什么 对其他细胞来说？发生坏死性下垂的细胞会影响周围的细胞。在这里，我们将询问ESCRT III是如何 对死亡细胞吞噬的影响，死亡延迟如何支持基因表达和蛋白质 产生，并允许对与死亡细胞相关的抗原进行适应性免疫。 完成我们的目标将促进我们对坏死性下垂是如何调节的，以及如何 它可以用于针对癌症和其他病理疾病的治疗。

Abstract Necroptosis is a form of cell death, inhibited by the FADD-caspase-8-FLIP complex, in which the interaction of two protein kinases, RIPK1 and RIPK3, interact to promote or inhibit necrotic death of a cell. Activated RIPK3 phosphorylates the pseudokinase MLKL, which mediates necroptosis through action at the plasma membrane. We have probed the late events that surround MLKL activation and have found that prior to loss of plasma membrane integrity, cell death is reversible. We have implicated the ESCRT III pathway, which functions in endosomal trafficking, cytokinesis, and exocytosis, in counter-acting the membrane effects of active MLKL. This results in the release of “bubbles” of damaged plasma membrane from the cell surface, sustaining cell survival. Our central hypothesis, upon which this application is based, is that necroptosis proceeds via the activation of MLKL and its action at the plasma membrane, and the regulation of this late process allows other events, such as protein expression induced by the necroptotic stimulus to manifest and thereby impact on the effects of the dying cell on the body. Based on this hypothesis, we will ask: 1. How do the late events in necroptosis regulate cell death and survival? Here we will identify the components of ESCRT III responsible for bubble formation after MLKL activation, and their role in “resuscitation” when active MLKL is subsequently disrupted. We will determine how ESCRT III functions to inhibit necroptosis in response to inducers without caspase inhibition, and explore if RIPK3 activity, independently of MLKL, may regulate the ESCRT III pathway. 2. Are there non-canonical pathways to MLKL activation and necroptosis? While several inducers of necroptosis have been identified, we propose that there are others that are less explored. We have identified DNA damage as an inducer of necroptosis, and will explore how this occurs. Further, we have found agents that appear to activate MLKL and necroptosis independently of RIPK3, and will determine how this non-canonical activation occurs. 3. What are the consequences of the late events in necroptosis for other cells? Cells undergoing necroptosis affect surrounding cells. Here we will ask how ESCRT III impacts on engulfment of the dying cell, how the delay in death supports gene expression and protein production, and allows engagement of adaptive immunity to antigens associated with the dying cell. Completion of our goals will advance our fundamental understanding of how necroptosis is regulated, and how it can be exploited in therapies directed to cancer and other pathological conditions.