Regulation of Inducible Nitric Oxide Synthase Asthma

诱导型一氧化氮合酶(INOS)诱导的一氧化氮(NO)的过度产生与 哮喘呼吸道炎症的发病机制。这项研究的长期目标是了解 调节iNOS活性，并设计新的方法来调节它。尽管很多人都知道 影响iNOS合成和催化活性的因素，人们对其细胞调控知之甚少。我们 最近发现iNOS是通过泛素-蛋白酶体途径降解的。它的特殊性 泛量化系统主要由特定的泛素连接酶(E3)提供，该酶识别和 结合到目标蛋白上。我们的初步数据确定了一种含有F-box的蛋白质；我们将其命名为UBLinos 这很可能是iNOS的E3泛素连接酶。其他初步数据表明，细胞 通过对诱导型一氧化氮合酶的时空调控来调控一氧化氮的合成。这些机制包括相对 诱导型一氧化氮合酶的快速转换和聚集到核周的位置，我们称之为“生理学” 此外，我们最近发现一类特定的咪唑类化合物 通过共翻译抑制iNOS组装而起作用。因此，使用这些化合物代表了一种 这是一种特异而有效的抑制iNOS产生的方法。 我们建议检验以下假设：a)含有F-box的蛋白(UBLinos)是E3连接酶 INOS，在iNOS细胞调控中起着核心作用。B)细胞的时间和空间调节 INOS的表达在观察到的与呼吸道炎症相关的iNOS上调中起着关键作用。 诱导型一氧化氮合酶翻译抑制剂降低实验性小鼠诱导型一氧化氮合酶水平并减轻炎症反应 哮喘模型。为了验证这些假设，我们提出了以下具体目标的研究：1) UBLinos对iNOS调控机制的确定。2)阐明了细胞的时态和 诱导型一氧化氮合酶在呼吸道炎症中的空间调控3)评价诱导型一氧化氮合酶翻译抑制剂在 通过在哮喘小鼠模型上测试它们的效果来测试它们对实验性哮喘的影响。上述研究将于#年进行。 培养的细胞以及从正常人、哮喘患者和 哮喘的小鼠模型。这些研究的结果将加深我们对监管的理解 诱导型一氧化氮合酶在呼吸道炎症中的合成，为制定治疗策略奠定基础。 在这种紊乱中调节iNOS。

Overproduction of nitric oxide (NO) by inducible nitric oxide synthase (iNOS) has been implicated in the pathogenesis of airway inflammation of asthma. The long-term goal of this research is to understand the regulation of iNOS activity and to devise novel methods to regulate it. Although much is known about factors affecting the synthesis and catalytic activity of iNOS, little is known about its cellular regulation. We have recently shown that iNOS is degraded through the ubiquitin-proteasome pathway. The specificity of the ubiqutination system is mainly provided by the specific ubiquitin ligase enzyme (E3) that recognizes and binds to the target protein. Our preliminary data identified an F-box-containing protein; we termed UBLinos that is a likely candidate to be the E3 ubiquitin ligase for iNOS. Additional preliminary data suggest that cells regulate NO synthesis by temporal and spatial regulation of iNOS. These mechanisms include a relatively rapid rate of iNOS turnover and sequestration of iNOS to a perinuclear location we termed the "physiologic aggresome." Furthermore, we have recently discovered that a specific class of imidazole like compounds acts by co-translational inhibition of iNOS assembly. Thus, the use of these compounds represents a specific and an efficient method to inhibit iNOS production. We propose to test the following hypotheses: A) An F-box-containing protein (UBLinos) is the E3 ligase for iNOS and it plays a central role in iNOS cellular regulation. B) The cellular temporal and spatial regulation of iNOS plays a critical role in the observed upregulation of iNOS associated with airway inflammation. iNOS translational inhibitors will reduce iNOS levels and ameliorate inflammation in experimental mouse models of asthma. To test these hypotheses we propose studies with the following specific aims: 1) Determination of mechanisms of iNOS regulation by UBLinos. 2) Elucidation of the cellular temporal and spatial regulation of iNOS in airway inflammation. 3) Evaluation of the use of iNOS translational inhibitors in experimental asthma by testing their effects in mouse models of asthma. The above studies will be done in cultured cells as well as in primary cells obtained from normal subjects, patients with asthma and from mouse models of asthma. The results of these studies will enhance our understanding of the regulation of NO synthesis by iNOS in airway inflammation and lay the groundwork for therapeutic strategies aimed at regulating iNOS in such disorder.