Heterotrimeric G-protein regulation of neurotransmission in C. elegans

通过异源三聚体GTP酶(G蛋白)的信号激活细胞效应酶和离子 调节细胞生长、有丝分裂和感官的通道。G蛋白偶联受体是 是真核生物中最常用的信号转导分子之一，并且是大约一半的 处方药物。土壤线虫，秀丽线虫，提供了许多实验性的 优势研究异源三聚体G蛋白信号的保守特征。在胚胎中，G- 蛋白质调节微管力，微管力控制细胞不对称分裂过程中有丝分裂纺锤体的定位。 在成年人中，G蛋白通过改变突触活性和 肌肉收缩的频率。这些行为为研究提供了方便和定量的分析方法。 遗传上的细胞内和细胞间信号传递。 通过三条独立的实验路线，我将研究分子和细胞的后果 通过两种G蛋白，EGL-30(Gctq)和GoA-1(GA0)传递信号。活化的EGL-30与EGL- 8，磷脂酰肌醇(4，5)二磷酸(PlPz)特异性磷脂酶C(P)家族成员。 PIP的水解？通过EGL-8释放第二信使1，4，5-三磷酸肌醇(IP3)和1，2-三磷酸 甘油二酯(DAG)。相比之下，激活的GOA-1的直接效应尚不清楚。为了找到这些，我会 分离抑制GOA-1信号表型过度激活的突变体。已经有一位抑制者 已经被科勒实验室的成员绘制了图谱，我将绘制更多的抑制子突变图，确定如何 编码的因子调节信号，并测试它们是否对GOA-1起直接效应作用。第二，到 研究GoA-1如何拮抗EGL-30信号调节突触活动，我将可视化EGL-8的活性 在体内使用已建立的钙和特定磷脂的荧光传感器。我将测试突变是如何 损害或刺激抑制性GOA-1信号会影响这些指标的行为和分布。 这些原位生化实验将揭示信号通路的细胞生物学后果。 从基因上定义的。最后，为了找到其表达受信号调节的基因，我将比较基因 微阵列分析GoA-1和EGL-30突变体的表达谱。 这项研究将阐明神经系统如何控制肌肉收缩的频率。 由于这些功能在人类大脑癫痫发作、肌肉震颤和神经退行性变期间受到干扰 对于帕金森氏症等疾病，这些研究应该为靶向治疗提供合理的基础。

Signaling through heterotrimeric GTPases (G-proteins) activates cellular effector enzymes and ion channels to regulate cell growth, mitotic division, and sensory perception. G-protein coupled receptors are among the most commonly used signal transducers in eukaryotes and are the targets of about half of prescribed Pharmaceuticals. The soil nematode, Caenorhabditis elegans, offers many experimental advantages to investigate the conserved features of heterotrimeric G-protein signaling. In embryos, G- proteins regulate microtubule forces that control mitotic spindle positioning during asymmetric cell division. In adults, G-proteins coordinate animal feeding, motility, and other behaviors by altering synaptic activity and the frequency of muscle contractions. These behaviors offer convenient and quantitative assays to study intracellular and intercellular signaling genetically. Through three independent lines of experimentation, I will study the molecular and cellular consequences of signaling through two G-proteins, EGL-30 (Gctq) and GOA-1 (Ga0). Activated EGL-30 interacts with EGL- 8, the phosphatidylinositol (4,5) bisphosphate (PlPz)-specific Phospholipase C (p) family member. Hydrolysis of PIP? by EGL-8 releases the second messengers inositol 1,4,5-triphosphate (IP3) and 1,2- diacylglycerol (DAG). In contrast, direct effectors of activated GOA-1 remain unknown. To find these, I will isolate mutants that suppress hyperactivated GOA-1 signaling phenotypes. One suppressor has already been mapped by members of the Koelle lab, and I will map additional suppressor mutations, determine how the encoded factors regulate signaling, and test whether they act as direct effectors for GOA-1. Second, to study how GOA-1 antagonizes EGL-30 signaling to modulate synaptic activity, I will visualize EGL-8 activity in vivo using established fluorescent sensors of Ca+2 and specific phospholipids. I will test how mutations that impair or stimulate inhibitory GOA-1 signaling affect the behavior and distribution of these indicators. These in situ biochemical experiments will reveal the cell biological consequences of the signaling pathways defined genetically. Finally, to find genes whose expression is regulated by signaling, I will compare gene expression profiles in goa-1 and egl-30 mutants by microarrayanalysis. This research will shed light on how the nervous system controls the frequency of muscle contractions. As these functions are perturbed during human brain seizures, muscle tremors, and neurodegenerative diseases such as Parkinson's, these studies should inform a rational basis for targeted therapies.