Genetic Analysis of Ras and G Protein Function

描述(由申请人提供)：复杂功能的执行及其在疾病中的分解涉及动物的遗传和环境之间的相互作用。该提案的总体目标是揭示RAS家族GTP酶如何影响控制突触可塑性的信号网络，以及“丰富的环境”(EE)如何改变这些网络，以改变青春期小鼠以及跨世代突触可塑性的诱导方式。这项建议的一个重点是GRF1和GRF2，它们形成了一系列多催化的、钙刺激的鸟嘌呤核苷酸交换因子，有可能激活RAS和RAC GTP酶。尽管有这些相似之处，我们发现GRF1和GRF2在青春期早期就促进了由NMDA型谷氨酸受体(NMDA-Rs)诱导的相反形式的突触可塑性。GRF1促进长期抑制(LTD)，而GRF2促进长期增强(LTP)，至少部分是因为它们调节不同的MAP激酶。下面概述的实验结合了遗传、生化和电生理学研究，以揭示GRF1和GRF2如何对不同的上游信号做出反应，以及它们的RAS和RAC激活区域的下游信号如何在海马体中进行差异调控。这些实验将为如何在神经元信号转导中实现特异性提供新的见解。它们还将显著增加我们对LTP和LTD诱导的分子基础的理解。这些成熟的学习和记忆细胞模式的缺陷被认为是导致各种神经和精神健康障碍的原因。这项建议的第二个重点是环境刺激，包括接触新对象、加强社会化和自愿锻炼，特别是在青春期前，如何改变诱导LTP的方式。我们发现，青春期的丰富解锁了一条以前未知的潜在信号通路，该通路促进了小鼠的LTP，并挽救了GRF基因敲除小鼠的LTP缺陷和上下文恐惧记忆。更具戏剧性的是，我们发现，青春期前丰富的这些影响会在青春期传递给下一代。所描述的实验将使用多种方法来揭示这一新的EE门控信号通路如何促进LTP，以及EE如何解锁这一级联反应以影响突触的可塑性和跨代记忆。更好地理解环境对大脑功能的跨代影响，可能会揭示克服神经和心理健康障碍的新方法。 公共卫生相关性：尽管一个人的基因蓝图极大地影响了他/她对疾病的易感性，但很明显，一个人所处的环境会影响对该蓝图的阅读方式。我们的研究调查了RAS蛋白如何对大脑中调节学习和记忆的生化途径做出贡献。它还探索了刺激的“丰富的环境”，特别是在青春期前，如何改变正常动物的这些生化途径，并补偿RAS信号的遗传缺陷。值得注意的是，我们发现，幼年富集物不仅会影响直接接触它的动物，还会影响它们未来青春期的后代。充分了解环境对大脑功能的跨代影响，可能会揭示克服神经和心理健康障碍的新方法。

DESCRIPTION (provided by applicant): The execution of complex functions and their breakdown in disease involves the interplay between an animal's genetics and environment. The overall goals of this proposal are to reveal how Ras-family GTPases influence signaling networks that control synaptic plasticity, and how an "enriched environment" (EE) alters these networks to change the way synaptic plasticity is induced in adolescent mice and remarkably, across generations. One focus of this proposal is on GRF1 and GRF2, which form a family of multi-catalytic, calcium- stimulated, guanine nucleotide exchange factors that have the potential to activate both Ras and Rac GTPases. Despite these similarities, we found that GRF1 and GRF2 promote opposing forms of synaptic plasticity induced by NMDA-type glutamate receptors (NMDA-Rs) beginning at early adolescence. GRF1 promotes long-term depression (LTD), while GRF2 promotes long-term potentiation (LTP), at least in part, because they regulate different MAP kinases. The experiments outlined below combine genetic, biochemical and electrophysiological studies to reveal how GRF1 and GRF2 respond to different upstream signals, and how signaling downstream from their Ras- and Rac-activating domains is differentially regulated in the hippocampus. These experiments will add new insight into how specificity is achieved in neuronal signal transduction. They will also add significantly to our understanding of the molecular basis of LTP and LTD induction. Defects in these well-established cellular paradigms of learning and memory are thought to contribute to a variety of neurological and mental health disorders. A second focus of this proposal is how environmental stimulation, involving exposure to novel objects, enhanced socialization and voluntary exercise particularly during pre-adolescence, changes the way LTP is induced. We discovered that adolescent enrichment unlocks a previously unidentified latent signaling pathway that promotes LTP in mice and rescues defective LTP and contextual fear memory in GRF knockout mice. Even more dramatic is our finding that these effects of pre-adolescent enrichment are passed on to the next generation through their adolescence. The experiments described will use multiple approaches to reveal how this novel EE-gated signaling pathway promotes LTP, and how EE unlocks this cascade to affect synaptic plasticity and memory across generations. A better understanding of the trans-generational effects of the environment on brain function may reveal new approaches to overcome neurological and mental health disorders. PUBLIC HEALTH RELEVANCE: Although a person's genetic blueprint strongly contributes to his/her susceptibility to disease, it is clear that the environment in which one lives influences how that blueprint is read. Our study investigates how Ras proteins contribute to biochemical pathways in the brain that mediate learning and memory. It also explores how a stimulating "enriched environment", particularly during pre-adolescence, changes these biochemical pathways in normal animals, and compensates for a genetic defect in Ras signaling. Remarkably, we find that juvenile enrichment affects not only animals directly exposed to it, but also their future offspring through adolescence. A full understanding of the trans-generational effects of the environment on brain function may reveal new approaches to overcome neurological and mental health disorders.