Bitter and sweet taste receptor physiology in airway ciliated cells

项目总结 呼吸道细胞纤毛T2R苦味受体检测细菌酰基高丝氨酸内酯 还有喹诺酮类药物。这些T2Rs的激活会增加一氧化氮(NO)的产生，从而增加 纤毛击打和杀灭细菌。因此，T2Rs在呼吸道先天免疫中起着重要作用。 纤毛T2Rs功能降低的多态与革兰氏阴性菌增多相关 上呼吸道感染，对慢性鼻窦炎的易感性，以及更糟糕的手术结果 在鼻窦手术后。靶向T2R或增强其活性可能是一种补充或 抗生素的替代品。增强呼吸道或其他组织的内源性T2R介导的免疫 面对日益普遍的肺炎，器官可能是一种有用的抗病原体策略 耐抗生素的“超级细菌”。NO可以杀菌，也可以灭活病毒。 T2Rs还能激活巨噬细胞等其他组织和免疫细胞中NO的生成， 其中，T2R到NO信号调节吞噬作用。T2Rs激活血管内皮细胞(E)型 原代呼吸道细胞和巨噬细胞中的一氧化氮合酶(ENOS)。这条路 在许多细胞类型中似乎是T2R信号的标志，并调节 生理过程。了解T2Rs如何向eNOS发出信号对于理解 口外味觉感受器在许多组织中的信号。我们假设T2R到eNOS的信号 需要通过激活T2Rs下游的激酶来磷酸化eNOS。激活剂 与许多其他GPCR相比，T2Rs激活的通路是非常未知的。我们也 假设这一激酶的磷酸化被重要的细胞因子HSP90增强 伴侣蛋白参与了一氧化氮合酶的功能以及一些gpr信号转导。 在这个方案中，我们将研究T2R介导的一氧化氮合酶和 手术残留物对原代鼻腔上皮细胞产生一氧化氮的影响 材料，并在气液界面生长(目标1)。我们将使用NO生产的活细胞成像 和生物化学与表征良好的激酶药理抑制剂相结合 已知的激活eNOS的途径。我们还将使用具有良好特性的药理HSP90 HSP90是否调节T2R或eNOS定位于气道纤毛或功能的抑制剂 (钙信号或NO产生)。

PROJECT SUMMARY T2R bitter taste receptors in airway cell cilia detect bacterial acyl-homoserine lactones and quinolones. Activation of these T2Rs increases nitric oxide (NO) production to increase ciliary beating and kill bacteria. T2Rs thus play an important role in airway innate immunity. Polymorphisms that reduce the function of cilia T2Rs correlate with increased gram negative upper respiratory infections, susceptibility to chronic rhinosinusitis, and worse surgical outcomes after sinus surgery. Targeting T2Rs or enhancing their activity may be a complementary or alternative to antibiotics. Boosting endogenous T2R-mediated immunity in the airway or other organs may be a useful anti-pathogen strategy in the face of the increasing prevalence of antibiotic-resistant “superbugs.” NO can be bactericidal and also can inactivate viruses. T2Rs also activate NO production in other tissues and immune cells like macrophages, where T2R to NO signaling regulates phagocytosis. T2Rs activate the endothelial (e) form of nitric oxide synthase (eNOS) in both primary airway cells and macrophages. This pathway appears to be a hallmark of T2R signaling in many cell types and regulates multiple physiological processes. Understanding how T2Rs signal to eNOS is critical for understanding extraoral taste receptor signaling in many tissues. We hypothesize that T2R to eNOS signaling requires phosphorylation of eNOS by kinases activated downstream of the T2Rs. Kinase pathways activated by T2Rs are very unexplored compared with many other GPCRs. We also hypothesize that this kinase phosphorylation is enhanced by HSP90, an important cellular chaperone protein involved in NOS function as well as some GPCR signaling. In this proposal, we will investigate T2R-mediated phosphorylation of NOS and consequences for NO production in primary nasal epithelial cells isolated from residual surgical material and grown at air-liquid interface (Aim 1). We will use live cell imaging of NO production and biochemistry combined with well-characterized pharmacological inhibitors of kinase pathways known to activate eNOS. We will also use well characterized pharmacological HSP90 inhibitors to characterize if HSP90 regulates T2R or eNOS localization to airway cilia or function (calcium signaling or NO production).