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程序，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主要的细胞命运。凋亡是一个被广泛研究的细胞死亡过程，它需要半胱天冬酶蛋白酶，并伴有典型的形态学特征。令人惊讶的是，缺乏凋亡效应物的小鼠存活到成年，这增加了非凋亡细胞死亡可能在动物发育中起关键作用的可能性。因此，一个主要的未解决的问题是，是否存在其他发育细胞死亡途径，如果存在，是什么分子机制控制它们的执行。我们最近发现秀丽隐杆线虫雄性特异性连接细胞（LC）的死亡不是凋亡。LC死亡既没有凋亡，也没有自噬或坏死的形态学特征。相反，垂死的LC表现出明显的核膜压痕，染色质未凝聚，内质网和线粒体肿胀。重要的是，LC死亡独立于CED-3 caspase，所有其他caspase和所有其他已知的秀丽隐杆线虫凋亡蛋白，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。通过全基因组RNAi筛选促进LC死亡的基因，我们确定了两个基因，pqn-41，一个以前未知功能的基因，let-70，编码E2泛素偶联酶。任何一种基因的缺失都会阻碍细胞核的形成，但不会阻碍细胞器的膨胀。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。在脊椎动物神经系统中，LC死亡与非凋亡性发育细胞死亡具有惊人的超微结构相似性。一些观察结果也表明与多q诱导的神经变性有相似之处。与polyQ蛋白一样，PQN-41C富含谷氨酰胺，并形成盘绕式二级结构。像polyQ蛋白一样，PQN-41C在细胞中聚集。此外，polyQ疾病组织的超微结构研究显示了与LC死亡时类似的变化，包括核膜细裂和细胞器肿胀。在此，我们建议了解pqn-41和let- 70以及相关哺乳动物蛋白的作用机制，并寻求小分子筛选来鉴定LC死亡抑制剂。总之，这些研究不仅应该告诉我们LC死亡的过程，而且可能为polyq依赖性神经退行性疾病的病因学提供重要线索和试剂。

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.