SGER: Role of CREB in Regulating Neuropeptide Synthesis

Pi：Nancy Lynne WayneNSF SGER Proposal#031407标题：“Role of CREB in Regulating NeuroPepal Composal”神经肽负责控制生理和行为的不同方面，包括身体生长、饮食、性别等。这项建议的广泛目标是了解使用控制软体动物繁殖的神经元来控制神经肽激素合成的细胞机制。该实验室早期的研究表明，电刺激生殖神经内分泌袋细胞神经元(BCNs)可导致神经肽产卵激素(ELH)的大量分泌。ELH的可释放量是通过上调ELH的合成来补充的，而牺牲了BCNs中的大多数其他蛋白质。结果表明，电刺激BCNs刺激了一条新的蛋白质合成途径，该途径涉及ELH mRNA上的一个特殊密码，称为内部核糖体进入位点(IRES)。这类似于病毒在感染后控制细胞中蛋白质合成的方式，并且被认为不在细胞系统中发挥生理作用。本研究的目的是了解刺激BCNs是如何激活ELH mRNA上的IRES的。在早期工作的基础上，假设cAMP第二信使途径(通过cAMP反应元件结合蛋白或CREB)在电刺激对IRES激活的影响中发挥重要作用，从而导致ELH合成上调。为了验证这一假说，CREB的表达将被RNA抑制所阻止，并且将记录对电刺激ELH合成和IRES激活的影响。这项实验综合运用了分子、生化、电生理和光学成像技术。预计抑制CREB将阻止IRES的电激活和随后ELH合成的上调。这项工作是以一种全新的方式思考细胞系统如何调节蛋白质合成的前沿工作。

PI: Nancy Lynne WayneNSF SGER proposal #031407Title: "Role of CREB in regulating neuropeptide synthesis"Neuropeptides are responsible for controlling different aspects of physiology and behavior including body growth, eating, drinking, sex, etc. The broad goal of this proposal is to understand cellular mechanisms controlling synthesis of a neuropeptide hormone using neurons that control reproduction in the mollusk Aplysia. Earlier studies from this laboratory showed that electrical excitation of the reproductive neuroendocrine bag-cell neurons (BCNs) led to large amounts of secretion of the neuropeptide egg laying hormone (ELH). The releasable amount of ELH was replenished by upregulating ELH synthesis at the expense of most other proteins in the BCNs. It was revealed that electrical excitation of the BCNs stimulated a novel protein-synthetic pathway involving a special code on ELH mRNA called an internal ribosomal entry site (IRES). This is similar to how viruses control protein synthesis in cells following infection, and was not thought to play a physiological role in cellular systems. The aim of the proposed studies is to understand how stimulation of the BCNs activates the IRES on ELH mRNA. Based on earlier work, it is hypothesized that the cyclic AMP second messenger pathway (via cyclic AMP response-element binding protein, or CREB) is playing an important role in mediating the effects of electrical excitation on IRES activation, leading to upregulation of ELH synthesis. To test this hypothesis, expression of CREB will be obstructed by RNA inhibition, and the effects on electrical excitation of ELH synthesis and IRES activation will be documented. This experiment uses a combination of molecular, biochemical, electrophysiological, and optical imaging techniques. It is expected that inhibition of CREB will prevent electrical activation of the IRES and subsequent upregulation of ELH synthesis. This work is at the forefront of a completely new way of thinking about the way that cellular systems can regulate synthesis of proteins