Neuronal Cell Death Regulation in Drosophila

果蝇神经细胞死亡调节

• 批准号: 7338294
• 负责人: FLORENCE M DAVIDSON
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7338294
• 关键词: Neuronal; Cell; Death; Regulation; Drosophila

An important question in biology is what makes cells die. We are studying the degeneration and premature cell death of terminally differentiated photoreceptor neurons (PRNs) in the adult retina. In flies and humans, many point mutations in the photoreceptor protein rhodopsin dominantly trigger PRN apoptosis and loss of vision. In humans, the disease is called retinitis pigmentosa (RP). The phenotype in Drosophila is remarkably similar to that in humans. Among other things, not only do PRNs expressing the mutant opsins die, but also PRNs that express different opsins. The mechanism is thus at least partly non-cell-autonomous. Our goal is to determine the mechanism of neuronal cell death and degeneration. Projects: Current research and future plans are focused on three major questions:1. The relationship between neurodegeneration and apoptosisQuantitation of neurodegeneration in vivo has historically been challenging. Yet it is essential for genetic modifier studies (see below). We have taken advantage of unique attributes of the fly compound eye to quantitate degeneration in vivo. But in the process of validating the assay, we discovered early, pre-apoptotic stages of degeneration whose relationship to classic apoptosis is unclear. We are particularly interested in the activation of cell death, so this is fortuitious. However, these degenerated morphologies often persist for weeks before apoptotic morphologies appear, whereas classic apoptosis takes only hours from start to finish. Amazingly, lowered dosages of proapoptotic genes such as hid suppress even these early changes. This raises the question of whether neurodegeneration in RP is a slowed-down form of apoptosis, or a separate process coincidentally controlled by hid and followed by apoptosis. Having determined specific methods of detection of these stages, we are engaged in identifying the genes involved using microarray, molecular biological and genetic approaches. 2. The trigger for degeneration provided by the dominant rhodopsin mutationsIn cases where RP mutations cause opsin misfolding, the leading theory has been that apoptosis is triggered by overstimulation of the unfolded protein response (UPR), constituting a toxic-gain-of-function mechanism. However, our research now shows that the mutant proteins are in fact partial loss-of-function mutants. This implies that wild type rhodopsin actually plays a role in PRN maintenance and survival, not just visual signaling. While the UPR undoubtedly occurs, the evidence does not support an essential role for it in cell killing. The mechanism we propose unifies all the previously reported data on PRN homeostasis, including studies of the effects of photoactivation in other phototransduction -signaling-mutant backgrounds, and the requirement for rhodopsin in PRN development. Most significantly, the mechanism makes us shift focus from "How do mutant opsins kill PRNs?" to "How does wild type opsin promote PRN maintenance and survival?" Given the evolutionary conservation of heterotrimeric G protein signaling, we anticipate that our results will be relevant to the control of apoptosis in a variety of cells.3. The genetic pathway of degeneration and cell death activated by rhodopsin mutations.By genetic deficiency screening, we have identified suppressor and enhancer loci affecting opsin mutant induced degeneration. Identification of the responsible genes is a future focus. In addition, candidate gene screening indicates both similarities and dissimilarities in how cell death is controlled in adult PRNs vs. developing PRNs and mitotic cells. These factors include light, cell-death-regulatory genes such as hid, TRAF, caspases and dIAP1, and the ras mitogen-activated kinase pathway genes. However, the activating or suppressing activities of these genes are sometimes opposite to those in mitotic cells, indicating different or additional control mechanisms in the adult, terminally differentiated neurons.

生物学中的一个重要问题是是什么导致细胞死亡。我们正在研究成人视网膜中终末分化感光神经元（PRN）的退化和过早细胞死亡。在果蝇和人类中，光感受器蛋白视紫红质的许多点突变主要引发 PRN 细胞凋亡和视力丧失。在人类中，这种疾病被称为色素性视网膜炎（RP）。果蝇的表型与人类的表型非常相似。除此之外，不仅表达突变视蛋白的 PRN 会死亡，表达不同视蛋白的 PRN 也会死亡。因此，该机制至少部分是非细胞自主的。我们的目标是确定神经元细胞死亡和退化的机制。项目：当前的研究和未来的计划集中在三个主要问题：1。神经变性和细胞凋亡之间的关系体内神经变性的定量历来具有挑战性。然而，它对于基因修饰研究至关重要（见下文）。我们利用果蝇复眼的独特属性来定量体内退化。但在验证该测定的过程中，我们发现了早期的凋亡前变性阶段，其与典型细胞凋亡的关系尚不清楚。我们对细胞死亡的激活特别感兴趣，所以这是偶然的。然而，这些退化的形态在凋亡形态出现之前通常会持续数周，而经典的细胞凋亡从开始到结束只需要几个小时。令人惊讶的是，降低促凋亡基因（例如 hid）的剂量甚至可以抑制这些早期变化。这就提出了一个问题：RP 中的神经变性是否是一种减慢的细胞凋亡形式，或者是一个由 hid 同时控制并随后发生细胞凋亡的单独过程。在确定了检测这些阶段的具体方法后，我们正在利用微阵列、分子生物学和遗传学方法来鉴定所涉及的基因。 2. 显性视紫红质突变引发的退化在 RP 突变导致视蛋白错误折叠的情况下，主要理论是细胞凋亡是由未折叠蛋白反应 (UPR) 的过度刺激引发的，构成了毒性功能获得机制。然而，我们现在的研究表明，突变蛋白实际上是部分功能丧失的突变体。这意味着野生型视紫红质实际上在 PRN 维持和存活中发挥作用，而不仅仅是视觉信号传导。虽然 UPR 无疑会发生，但证据并不支持它在细胞杀伤中发挥重要作用。我们提出的机制统一了所有先前报道的关于 PRN 稳态的数据，包括对其他光转导信号突变背景中光激活效应的研究，以及 PRN 发育中视紫红质的需求。最重要的是，该机制使我们将焦点从“突变视蛋白如何杀死 PRN？”转移。到“野生型视蛋白如何促进 PRN 维持和存活？”鉴于异源三聚体G蛋白信号传导的进化保守性，我们预计我们的结果将与多种细胞中细胞凋亡的控制相关。3．视紫红质突变激活变性和细胞死亡的遗传途径。通过遗传缺陷筛选，我们鉴定了影响视蛋白突变体诱导变性的抑制子和增强子位点。识别相关基因是未来的重点。此外，候选基因筛选显示了成人 PRN 与发育中 PRN 和有丝分裂细胞的细胞死亡控制方式的相似之处和不同之处。这些因子包括光、细胞死亡调节基因，如 hid、TRAF、半胱天冬酶和 dIAP1，以及 ras 丝裂原激活激酶途径基因。然而，这些基因的激活或抑制活性有时与有丝分裂细胞中的相反，表明成年终末分化神经元中存在不同或额外的控制机制。