Regulation of CD23 Cleavage

项目摘要 免疫球蛋白介导的过敏性哮喘影响了1000万美国人。过敏性哮喘发作是由暴露于引起支气管收缩的过敏原引发的，通过使用β-2-肾上腺素能受体(Beta{2}AR)激动剂来松弛细支气管周围的平滑肌细胞，可以缓解这种情况。肺中的其他细胞表达β2受体，包括产生IgE的B淋巴细胞。高水平的IgE会转化为更严重的过敏性哮喘症状。我们的实验室已经报道，在激活的B细胞上，Beta{2}AR刺激会增加IgE转录的速度。β受体的天然配体是神经递质去甲肾上腺素，它也与过敏性哮喘发作的严重程度有关。如果IgE水平与哮喘反应的严重程度相关，如果B细胞上的Beta{2}AR刺激增加了IgE，那么确定介导这种增加的机制将是重要的。IgE水平受膜CD23(MCD23)和可溶性CD23(SCD23)裂解的调节，后者分别负向或正向调节IgE的产生。我们的实验室发现，在β受体刺激B细胞后，胞浆中总CD23和sCD23的表达增加，而mCD23的表达保持不变，提示调控mCD23切割的机制可能是通过刺激B细胞后激活的信号中间产物来实现的。将mCD23裂解为sCD23的酶属于A去整合素和金属蛋白水解酶(ADAMS)家族，当ADAMS被激活时，它的功能是增加IgE的正向可溶性调节因子，同时保持负膜调节因子的恒定。虽然ADAM10被报道为主要的脱落酶，但我们的初步数据显示，激活后B细胞中ADAMS 8、10和28的水平增加，但只有ADAMS 8和28的水平在伴随的Beta{2}AR刺激后进一步增加。在这项提议中概述的实验中，我们将检验这样的假设，即B细胞上的Beta{2}AR刺激增加了sCD23/mCD23的比率和随后通过调节特定ADAMS的水平和/或蛋白分解活性而产生的IgE水平。检验这一假说的意义在于，这些数据将阐明B细胞上的Beta{2}AR刺激参与调节IgE水平的机制，从而在B细胞中识别新的药物靶点，以防止激动剂活性过度产生IgE，而不干扰其他细胞上的激动剂活性，从而缓解过敏性哮喘症状。

Project Summary IgE-mediated allergic asthma affects 10 million Americans. An allergic asthma attack is triggered by exposure to allergens that cause bronchoconstriction, which is relieved by the administration of a beta-2-adrenergic receptor (Beta{2}AR) agonist to relax smooth muscle cells surrounding the bronchioles. Other cells in the lung express the Beta{2}AR, including B lymphocytes that produce IgE. High levels of IgE translate into more severe allergic asthma symptoms. Our laboratory has reported that Beta{2}AR stimulation on an activated B cell increases the rate of IgE transcription. The natural ligand for the Beta{2}AR is the neurotransmitter norepinephrine, which is also associated with the severity of allergic asthma attacks. If the level of IgE is associated with the severity of an asthmatic response and if Beta{2}AR stimulation on a B cell increases IgE, it will be important to identify the mechanism that mediates this increase. IgE levels are regulated by the cleavage of membrane CD23 (mCD23) to soluble CD23 (sCD23), which, respectively, regulate IgE production negatively or positively. Our laboratory determined that there was a Beta{2}AR-induced increase in total cytoplasmic CD23 and sCD23, while mCD23 remained constant, suggesting that the mechanism involved in the regulation of mCD23 cleavage may be targeted by signaling intermediates activated after Beta{2}AR stimulation on a B cell. The enzymes that cleave mCD23 to sCD23 belong to a family of A Disintegrin And Metalloproteinases (ADAMs), which, when active, function to increase the positive soluble regulator of IgE, while keeping the negative membrane regulator constant. Although ADAM10 is reported to be the primary sheddase, our preliminary data show that the levels of ADAMs 8,10, and 28 increase in a B cell after activation, but that only the levels of ADAMs 8 and 28 increase further after concomitant Beta{2}AR stimulation. In experiments outlined in this proposal, we will test the hypothesis that Beta{2}AR stimulation on a B cell increases the ratio of sCD23/mCD23 and the subsequent level of IgE produced by regulating either the level and/or proteolytic activity of specific ADAMs. The significance of testing this hypothesis is that the data will elucidate a mechanism by which Beta{2}AR stimulation on a B cell participates in regulating the level of IgE, thus identifying novel drug targets in a B cell that would prevent agonist activity to overproduce IgE, without interfering with agonist activity on other cells that would relieve allergic asthma symptoms.