Heterotrimeric G-protein regulation of neurotransmission in C. elegans

通过异源三聚体GTP酶（G蛋白）的信号传导激活细胞效应酶和离子 调节细胞生长、有丝分裂和感觉知觉的通道。G蛋白偶联受体是 是真核生物中最常用的信号转导子之一， 处方药。土壤线虫，秀丽隐杆线虫，提供了许多实验， 优势，以调查的保守特征的异源三聚体G蛋白信号。在胚胎中，G- 在不对称细胞分裂期间，蛋白质调节控制有丝分裂纺锤体定位的微管力。 在成年人中，G蛋白通过改变突触活动和神经活动来协调动物的进食、运动和其他行为。 肌肉收缩的频率这些行为提供了方便和定量的分析研究 细胞内和细胞间的信号传递。 通过三个独立的实验，我将研究分子和细胞的后果 通过两种G蛋白EGL-30（Gctq）和果阿-1（Ga 0）进行信号传导。活化的EGL-30与EGL-30相互作用。 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释放第二信使肌醇1，4，5-三磷酸（IP 3）和1，2- 甘油二酯（DAG）。相反，激活的果阿-1的直接效应子仍然未知。为了找到这些，我会 分离抑制过度活化果阿-1信号传导表型的突变体。一个抑制器已经 Koelle实验室的成员已经绘制了地图，我将绘制更多的抑制基因突变，确定如何 编码的因子调节信号传导，并测试它们是否作为果阿-1的直接效应物。二是 为了研究果阿-1如何拮抗EGL-30信号传导以调节突触活性，我将可视化EGL-8活性 使用Ca+2和特定磷脂的已建立的荧光传感器在体内进行。我将测试突变 损害或刺激抑制性果阿-1信号传导的细胞因子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 基因定义最后，为了找到其表达受信号传导调控的基因，我将比较基因表达， 通过微阵列分析在果阿-1和egl-30突变体中的表达谱。 这项研究将揭示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疾病期间受到干扰， 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.