G Protein Signaling in the C. elegans Nervous System

描述（由申请人提供）：我们试图了解神经递质如何通过异源三聚体G蛋白发出信号以调节神经元的活动。成瘾性药物激活G蛋白偶联受体，许多精神疾病是由于通过G蛋白的神经传递的改变，因此了解神经元中G蛋白信号传导的基本机制至关重要。Galpha-o是G蛋白偶联神经递质受体激活的主要脑G蛋白，但对其信号传导机制知之甚少。最近的一项发现是Galpha-o在分裂细胞中起作用以调节微管上的力，提出了Galpha-o也可能通过微管力作用于神经元以改变细胞结构的假设。Galpha-o信号抑制C.我们的第一个目标是用它作为研究Galpha-o信号机制的模型。因此，我们将鉴定产卵系统中产生和接收Galpha-o介导的信号的细胞。我们将分析三个新的基因，我们已经从遗传学上确定了这是必要的信号，其中一个我们已经克隆和编码TRP离子通道。我们将通过荧光标记野生型和Galpha-o-突变动物的产卵系统的神经过程和突触来检验Galpha-o通过细胞骨架改变神经元结构的假设。我们的第二个目标是利用Galpha-o信号的第二种模型。因此，我们将研究Galpha-o介导的5-羟色胺信号传导机制，5-羟色胺是一种与人类抑郁症有关的神经递质。我们正在筛选C.对血清素没有反应的优雅动物我们将完成这项筛选，克隆和分析一个新的5-羟色胺信号基因，我们已经通过这种方法确定。我们还将分析一组C. elegans血清素受体同系物。我们的第三个目标是鉴定和分析在Galpha-o下游起作用以介导其效应的分子。我们将完成一个基因筛选突变破坏神经递质信号下游的Galpha-o。使用这个屏幕和5-羟色胺屏幕，我们已经分离出五个突变，似乎破坏了Galpha-o介导的神经传递以及微管力在分裂细胞中的产生。这有力地支持了Galpha-o通过相同的机制对两种功能起作用的假设。通过上位性分析，其中三个突变确定了一个单一的基因，该基因似乎在Galpha-o的下游发挥作用。我们将克隆和分析这些突变所识别的基因。

DESCRIPTION (provided by applicant): We seek to understand how neurotransmitters signal through heterotrimeric G proteins to modulate the activities of neurons. Addictive drugs activate G protein-coupled receptors, and a number of mental diseases are due to alterations in neurotransmission through G proteins, so it is critical to understand the fundamental mechanisms of G protein signaling in neurons. Galpha-o is the major brain G protein activated by G protein-coupled neurotransmitter receptors, but little is known about its mechanism of signaling. A recent discovery is that Galpha-o acts in dividing cells to regulate force on microtubules, raising the hypothesis that Galpha-o may also act in neurons via microtubule force to modify cell structure. Galpha-o signaling inhibits egg-laying behavior in C. elegans, and our first aim is to use this as a model to study the Galpha-o signaling mechanism. Thus we will identify the cells of the egg-laying system that generate and receive Galpha-o-mediated signal(s). We will analyze three new genes we have genetically identified that are required for this signaling, one of which we have already cloned and shown to encode a TRP ion channel. We will test the hypothesis that Galpha-o acts via the cytoskeleton to alter the structure of neurons by fluorescently labeling the neural processes and synapses of the egg-laying system in wild-type and Galpha-o-mutant animals. Our second aim exploits a second model for Galpha-o signaling. Thus we will study the mechanism of Galpha-o-mediated signaling by serotonin, a neurotransmitter involved in depression in humans. We are screening for mutants of C. elegans that fail to respond to serotonin. We will complete this screen and clone and analyze a new serotonin signaling gene that we have already identified by this approach. We will also analyze the expression patterns and knockout phenotypes for a set of C. elegans serotonin receptor homologs. Our third aim is to identify and analyze the molecules that act downstream of Galpha-o to mediate its effects. We will complete a genetic screen for mutants that disrupt neurotransmitter signaling downstream of Galpha-o. Using this screen and the serotonin screen, we have already isolated five mutations that appear to disrupt both Galpha-o-mediated neurotransmission as well as microtubule force generation in dividing cells. This strongly supports the hypothesis that Galpha-o acts by the same mechanism for both functions. Three of the mutations identify a single gene that, by epistasis analysis, appears to function downstream of Galpha-o. We will clone and analyze the gene(s) identified by these mutations.