Control of nonapoptotic C. elegans cell death similar to neurodegeneration

描述(由申请人提供)：我们的长期目标是了解我们发现的一种新的非凋亡性发育细胞死亡计划，以及它与聚谷氨酰胺诱导的神经退行性疾病的关系。细胞死亡是后生动物发育过程中的主要细胞命运。细胞凋亡是一种被广泛研究的细胞死亡过程，需要半胱氨酸天冬氨酸蛋白酶，并伴随着典型的形态特征。令人惊讶的是，缺乏凋亡效应的小鼠可以存活到成年，这增加了非凋亡性细胞死亡可能在动物发育中发挥关键作用的可能性。因此，一个尚未解决的主要问题是，是否存在可供选择的发育细胞死亡途径，如果存在，控制其执行的分子机制是什么。我们最近发现，线虫雄性特异性连接细胞(LC)的死亡不是凋亡的。LC死亡既不具有凋亡性，也不具有自噬或坏死的形态特征。相反，垂死的LC显示出明显的核膜凹陷(皱缩)、未浓缩的染色质以及内质网和线粒体的肿胀。重要的是，LC死亡不依赖于CED-3 caspase、所有其他caspase和所有其他已知的线虫凋亡蛋白，包括CED-4/APAF-1、CED-9/Bcl-2家族、EGL-1和CED-13 BH3结构域蛋白。这些令人兴奋的发现表明，LC死亡必须通过一种新的机制发生。从全基因组RNAi筛选促进LC死亡的基因中，我们发现了两个基因，PQN-41，一个以前未知的功能基因，以及LET-70，编码一个E2泛素结合酶。任何一种基因的丢失都会阻止核皱缩，但不会阻止细胞器的肿胀。PQN-41C转录本和LET-70促进LC死亡，并且只有在细胞死亡开始时才在LC中表达。LC死亡表现出与脊椎动物神经系统中非凋亡性发育细胞死亡显著的超微结构相似之处。一些观察也表明，这与多聚Q诱导的神经退行性变有相似之处。与PolyQ蛋白一样，PQN-41C富含谷氨酰胺，形成卷曲的二级结构。像多聚Q蛋白一样，PQN-41C聚集在细胞中。此外，多Q病组织的超微结构研究揭示了与LC死亡时类似的变化，包括核膜皱缩和细胞器肿胀。在这里，我们建议了解PQN-41和LET-70的作用机制，以及相关的哺乳动物蛋白，并寻求小分子筛选来识别LC死亡的抑制剂。总之，这些研究不仅应该让我们了解LC的死亡过程，而且可能会为多聚Q依赖型神经退行性疾病的病因学提供重要的线索和试剂。

DESCRIPTION (provided by applicant): Our long-term goal is to understand a novel nonapoptotic developmental cell death program we discovered, and its relationship to polyglutamine-induced neurodegenerative disease. Cell death is a major cell fate during metazoan development. Apoptosis, an extensively studied cell death process, requires caspase proteases and is accompanied by a stereotypical morphological signature. Surprisingly, mice lacking apoptotic effectors survive to adulthood, raising the possibility that non- apoptotic cell death may play key roles in animal development. Thus, a major unsolved question is whether alternative developmental cell death pathways exist, and if so, what molecular mechanisms govern their execution. We recently discovered that the death of the C. elegans male-specific linker cell (LC) is not apoptotic. LC death has neither apoptotic, nor autophagic or necrotic morphological features. Instead, the dying LC displays pronounced indentation (crenellation) of the nuclear envelope, uncondensed chromatin, and swelling of the endoplasmic reticulum and mitochondria. Importantly, LC death is independent of CED-3 caspase, all other caspases, and all other known C. elegans apoptotic proteins, including CED-4/Apaf-1, CED-9/Bcl-2 family, and EGL-1 and CED-13 BH3-domain-only proteins. These exciting findings demonstrate that LC death must occur through a novel mechanism. From a genome-wide RNAi screen for genes promoting LC death we identified two genes, pqn-41, a gene of previously unknown function, and let-70, encoding an E2 ubiquitin conjugating enzyme. Loss of either gene blocks nuclear crenellation, but not organelle swelling. The pqn-41C transcript as well as let-70 promote LC death, and are expressed in the LC only as cell death is initiated. LC death displays striking ultrastructural similarities to nonapoptotic developmental cell death in the vertebrate nervous system. Several observations also suggest similarities to polyQ-induced neurodegeneration. Like polyQ proteins, PQN-41C is highly glutamine rich, and forms coiled-coil secondary structures. Like polyQ proteins, PQN-41C aggregates in cells. Furthermore, ultrastructural studies of polyQ disease tissue reveal changes similar to those seen during LC death, including nuclear envelope crenellation and organelle swelling. Here we propose to understand the mechanisms of action of pqn-41 and let- 70, and related mammalian proteins, and to pursue a small molecule screen to identify inhibitors of LC death. Together, these studies should not only inform us about the process of LC death, but may yield important clues and reagents as to the etiology of polyQ-dependent neurodegenerative disease.