CYCLIC NUCLEOTIDE PHOSPHODIESTERASES IN OLFACTION

嗅觉中的环核苷酸磷酸二酯酶

• 批准号: 2518093
• 负责人: JAMES A. CHERRY
• 金额: $ 12.79万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-15 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2518093
• 关键词: 3'5' cyclic nucleotide phosphodiesterase; RNase protection assay; behavior test; biological signal transduction; calmodulin; chemoreceptors; cyclic AMP; developmental genetics; developmental neurobiology; embryo /fetus culture; experimental brain lesion; gene expression; gene targeting; immunocytochemistry; in situ hybridization; laboratory mouse; mammalian embryology; molecular cloning; nervous system regeneration; neurogenesis; neurogenetics; olfactions; olfactory lobe; respiratory epithelium; restriction mapping; sensory discrimination

The long-range goals of this research program are to define the fundamental cellular mechanisms involved in olfactory perception. A critical and active component of the molecular pathway underlying odorant transduction is the second messenger molecule cAMP, which has been shown to increase rapidly in olfactory sensory neurons upon odor stimulation. The focus of this proposal is on the enzymes that degrade these molecules- - cyclic nucleotide phosphodiesterases (PDEs)-- and are therefore well- situated to act as modulators of the olfactory signals that are ultimately transmitted to the brain. In mammals one of these enzymes is a homolog of the Drosophila dunce gene, which codes for a cAMP-specific PDE. This gene has recently been cloned in the mouse and shown to be abundantly and preferentially expressed in olfactory sensory neurons. A series of studies are proposed to extend this initial discovery. First, the developmental onset of the dunce homolog, called mPDE2, will be established during early formation of the olfactory system. Regulation of this enzyme will then be examined during degeneration and subsequent regeneration of the olfactory mucosa following unilateral removal of the olfactory bulb, which is the target of the sensory neuron axons. Techniques in molecular genetics will then be used to begin addressing basic questions concerning the role of this PDE in olfaction. First, the gene structure of mPDE2 will be studied, including investigations into the protein and/or mRNA heterogeneity in olfactory tissue. Then the mPDE2 gene will be disrupted, or knocked out, using homologous recombination in embryonic stem cells. Once the mice lacking a functional mPDE2 gene are obtained they will be thoroughly analyzed morphologically, physiologically, and behaviorally for any phenotypic abnormalities, particularly with respect to potential deficiencies associated with olfactory function. Finally, to address more completely the issue of PDE function in olfaction, it will be necessary to characterize the other major PDE found in sensory neurons-- a Ca2+ and calmodulin-activatible PDE, or CaM PDE. Therefore, identification of the CaM PDEs expressed in the olfactory system by molecular cloning is proposed. It is expected that these experiments will contribute important information about the role of cyclic nucleotide PDEs in olfaction.

这项研究计划的长期目标是确定 嗅觉的基本细胞机制。 一 气味物质分子途径的关键和活性成分 转导是第二信使分子cAMP，这已被证明是 嗅觉神经元在气味刺激后迅速增加。 这项提案的重点是降解这些分子的酶- - 环核苷酸磷酸二酯酶（PDE）-因此很好- 作为嗅觉信号的调节器， 传递到大脑。 在哺乳动物中，其中一种酶是 Drosophila dunce基因，编码cAMP特异性PDE。 该基因 最近被克隆到老鼠身上， 优先在嗅觉神经元中表达。 一系列 有人建议进行研究来扩展这一初步发现。 一是 一种叫做mPDE 2的傻瓜同系物的发育开始， 在嗅觉系统的早期形成过程中建立的。 调控 然后在变性和随后的 嗅粘膜的再生后，单侧切除的 嗅球，这是感觉神经元轴突的目标。 分子遗传学技术将被用于开始解决 关于这个PDE在嗅觉中的作用的基本问题。 一是 将研究mPDE 2的基因结构，包括对 蛋白质和/或mRNA的异质性。 然后mPDE 2 基因将被破坏，或敲除，使用同源重组， 胚胎干细胞 一旦缺乏功能性mPDE 2基因的小鼠， 获得它们将被彻底地形态学分析， 生理上和行为上的任何表型异常， 特别是关于与以下方面相关的潜在缺陷： 嗅觉功能 最后，为了更全面地解决PDE问题， 功能的嗅觉，这将是必要的特点，其他 在感觉神经元中发现的主要PDE-一种Ca 2+和钙调素激活的 PDE或CaM PDE。 因此，鉴定表达于细胞中的CaM PDE， 提出了通过分子克隆研究嗅觉系统的方法。 预计 这些实验将提供重要的信息， 环核苷酸PDE在嗅觉中的作用。