PHOSPHORYLATION OF CILIARY TARGET PROTEINS

描述（由申请人提供）：本项目的长期目标是了解调节气道疾病中粘膜纤毛清除及其功能障碍的细胞和分子机制。 最近在气道上皮细胞顶端发现透明质酸及其对纤毛功能的调节作用，使得透明质酸成为除ATP之外的第二种已知的内源性调节剂，调节气道腔侧的人纤毛搏动频率（CBF）。 因此，透明质酸必须在宿主防御中发挥重要作用。 初步数据表明，透明质酸是由上皮细胞以需要降解的形式产生的，以刺激CBF。 羟考酮可在气道中被活性氧/氮物质降解。 因此，在气道损伤期间，预期透明质酸通过未知的途径在纤毛细胞的顶端通过RHAMM（透明质酸介导的运动性或CD 168的受体）去活化并开始信号传导以刺激CBF。 已经提出在其他系统中的RHAMM信号传导通过透明质酸介导的与生长因子受体的关联起作用。 罗恩是HGF家族的一种受体，表达于人气道上皮细胞的顶端，可介导透明质酸对CBF的刺激作用。 因此，该提议将检验透明质酸在其固定的浅表上皮的顶膜中组装的总体假设。 在透明质酸被氧化剂解聚后，通过RHAMM的信号传导刺激CBF，进而刺激粘膜纤毛清除。 具体目标是：1）检验透明质酸滞留在表面的假设，透明质酸滞留是通过附着在上皮顶端的合成酶上或通过与RHAMM相互作用而发生的; 2）检验只有中等至低分子大小的透明质酸分子刺激CBF的假设; 3）检验内源性透明质酸通过氧化剂原位解聚增加CBF和粘液转运速率的假设;和4）检验经由RHAMM的信号传导通过透明质酸介导的RHAMM与生长因子受体（可能是罗恩）的结合而增加CBF和粘液转运速率的假设。这些实验的结果将为我们理解气道生物学提供新的机制见解，因为它涉及透明质酸，纤毛跳动，粘膜纤毛清除及其在气道中的相互作用，并可以确定粘膜纤毛功能障碍的新治疗靶点。

DESCRIPTION (provided by applicant): The long-term goal of this project is to understand the cellular and molecular mechanisms that regulate mucociliary clearance and their dysfunction in airway diseases. The recent discovery of hyaluronan at the apex of airway epithelial cells and its regulation of ciliary function makes hyaluronan only the second known endogenously produced regulator, in addition to ATP, of human ciliary beat frequency (CBF) at the luminal side of the airway. Thus, hyaluronan must play an important role in host defense. Preliminary data suggest that hyaluronan is made by the epithelium in a form that requires degradation to stimulate CBF. Hyaluronan can be degraded in the airways by reactive oxygen/nitrogen species. Thus, during times of airway insults, hyaluronan is expected to depolymerize and start signaling via RHAMM (receptor for hyaluronic acid mediated motility or CD 168) at the apex of ciliated cells through an unknown pathway to stimulate CBF. RHAMM signaling in other systems has been proposed to work through hyaluronan-mediated association with growth factor receptors. RON, a receptor of the HGF family that can regulate CBF, is expressed at the apex of human airway epithelial cells and could mediate hyaluronan's stimulation of CBF. Thus, this proposal will test the overall hypothesis that hyaluronan is assembled in the apical membrane of the superficial epithelium where it is immobilized. Upon hyaluronan depolymerization by oxidants, signaling through RHAMM stimulates CBF and in turn mucociliary clearance. The specific aims are: 1) to test the hypothesis that hyaluronan retention at the surface occurs either by attachment to its synthase at the epithelial apex or by interaction with RHAMM; 2) to test the hypothesis that only mid to low molecular sized molecules of hyaluronan stimulate CBF; 3) to test the hypothesis that in situ depolymerization of endogenous hyaluronan by oxidants increases CBF and mucus transport rates; and 4) to test the hypothesis that signaling via RHAMM increases CBF and mucus transport rates by hyaluronan-mediated association of RHAMM with a growth factor receptor, possibly RON. The results of these experiments will provide new mechanistic insights into our understanding of airway biology as it pertains to hyaluronan, ciliary beating, mucociliary clearance, and their interactions in the airways, and could identify novel therapeutic targets for mucociliary dysfunction.