OMP: Role in Olfactory Signal Detection and Transduction

描述(由申请人提供)：钙在嗅觉感受器神经元(ON)的信号转导级联中扮演许多角色。钙离子通过cAMP门控通道进入，反应气味配体受体激活的G蛋白介导的腺酰环化酶活性的增加。Ca~(2+)内流是哺乳动物嗅觉化学感觉转导的关键步骤，它刺激钙激活的氯离子通道来放大转导电流。Ca2+是 也是适应、脱敏、恢复和转导-翻译耦合的调节器。为了理解ORN中的这些事件，人们付出了大量的努力。在Orns中维持稳定的细胞内钙水平的下游事件受到的关注要少得多。我们对嗅觉标记蛋白(OMP)缺失小鼠表型的鉴定 在之前的资金周期中，使用电生理、生化和行为指标使我们假设OMP无效表型是由于调节细胞内钙离子水平的缺陷所致。具体地说，我们假设OMP参与调节ONS中钙离子排出过程的活性。OMP在成熟脊椎动物中表达的特异性，其序列在所有脊椎动物中的保守性，以及OMP缺失的小鼠表型ALL 提示OMP对嗅觉功能有重要作用。我们最近观察到OMP缺失的小鼠在刺激后树突状细胞结节内的钙动力学发生了显著的变化。参与钙外排的基因有Na+/Ca~(2+)交换器(NCX)和Na~+/Ca~(2+)-K~+交换器(NCKX)。三个基因代表这些基因中的每一个，以及以细胞和组织特有的模式表达的大量mRNA剪接变体。免疫细胞化学和电生理报告表明，这些交换器在维持细胞内ORN水平方面起着重要作用。这些报告，再加上我们的新数据，即OMP调节Ca~(2+)排出，进一步支持了我们的假设和表征ONS中Ca~(2+)排出过程的分子和生化基础的重要性。这是我们在这项提议中所解决的知识上的一个重大差距。 我们将(1)-鉴定NCX和NCKX基因在嗅觉组织中的表达，特别是在嗅觉组织发育期间和对病变的反应中；(2)-确定参与OMP介导的钙水平调节的特定NCX或NCKX蛋白的部分；(3)-确定这一过程中其他参与者的性质和功能，例如最近发现的与OMP结合的Bex蛋白。这些研究将为这些机制提供新的见解。 在ONS中，细胞内的钙离子水平被调节，并可能为嗅觉障碍或嗅觉障碍的治疗提供化疗机会。

DESCRIPTION (provided by applicant): Calcium plays many roles in the signal-transduction cascade in olfactory receptor neurons (ORNs). Ca2+ enters through cAMP-gated channels in response to odor-ligand receptor activation of G-protein-mediated increases in adenylyl cyclase activity. Ca2+ entry is a key step in mammalian olfactory chemosensory transduction and stimulates a Ca2+-activated chloride channel to amplify the transduction current. Ca2+ is also a regulator of adaptation, desensitization, recovery and transduction-translation coupling. Much effort has been directed towards understanding these events in the ORN. Much less attention has been paid to downstream events that maintain stable intracellular Ca2+ levels in ORNs. Our characterization of the olfactory marker protein (OMP)-null mouse phenotype using electrophysiological, biochemical and behavioral measures in the previous funding cycle have led us to hypothesize that the OMP-null phenotype is due to a defect in regulating intracellular Ca2+ levels. Specifically, we hypothesize that OMP participates in regulating the activity of Ca2+ extrusion processes in ORNs. The specificity of OMP expression in mature ORNs, its sequence conservation across all vertebrates, and the OMP-null mouse phenotype all indicate OMP is important to olfactory function. We recently observed a significant alteration in Ca2+ kinetics in ORIN dendritic knobs in OMP-null mice following stimulation. Several genes participate in Ca2+ extrusion including the Na+/Ca2+ exchangers (NCX), and the Na+/Ca2+-K+ exchangers (NCKX). Three genes represent each of these and numerous mRNA splice variants expressed in cell- and tissue-specific patterns. Immunocytochemical and electrophysiological reports suggest that these exchangers are important in maintaining intracellular ORN levels. These reports, coupled with our new data that OMP modulates Ca2+ extrusion further supports our hypothesis and the importance of characterizing the molecular and biochemical bases of Ca2+ extrusion processes in ORNs. This is a significant gap in our knowledge that this proposal addresses. We will (1)-characterize NCX and NCKX gene expression in olfactory tissue, and especially in ORNs, during development and in response to lesions; (2)-identify that part of the specific NCX or NCKX protein that participates in the OMP-mediated regulation of Ca levels; (3)-determine the nature and function of other participants in this process e.g. the recently discovered Bex proteins that bind to OMP. These studies will provide novel insights to the mechanisms by which intracellular Ca2+ levels are regulated in ORNs and may generate chemotherapeutic opportunities for treatment of anosmias or dysosmias.