Prohormone processing: NPY/Catestatin Peptide Production

神经肽Y（NPY）和catestatin肽由肾上腺髓质嗜铬细胞和交感神经分泌，用于调节血压。NPY作为直接的血管收缩剂，catestatin作为自分泌调节剂，抑制尼古丁刺激的儿茶酚胺释放。原发性高血压患者中NPY升高和catestatin降低提示其作为神经效应物参与血压调节。重要的是，了解负责将其各自的激素前体转化为活性NPY和catestatin的主要蛋白水解酶对于理解控制血压的调节机制至关重要。神经肽Y和睾丸素的激素原前体、神经肽Y原和嗜铬粒蛋白A（CgA）分别在肾上腺髓质的分泌囊泡（称为嗜铬颗粒）内进行蛋白水解加工。我们已经确定了分泌囊泡组织蛋白酶L，以前称为？激素原巯基蛋白酶？(PTP)作为神经肽Y原和CgA的关键加工酶。此外，本项目还发现了一种新的内源性丝氨酸蛋白酶抑制剂，即内桥蛋白2，它能抑制分泌囊泡组织蛋白酶L。此外，枯草杆菌蛋白酶样 分泌囊泡中的PC 1和PC 2蛋白酶也可能参与pro-NPY和CgA的加工。基于这些新的发现，项目3的目标将是评估分泌囊泡组织蛋白酶L和endopin 2与PC 1和PC 2相比在产生调节血压的NPY和catestatin神经肽中的作用。本项目将测试的假设，分泌囊泡组织蛋白酶L可能是一个主要的加工酶的神经肽Y和catestatin相比，PC 1和PC 2酶。我们的新研究结果支持组织蛋白酶L在分泌囊泡中对NPY原和CgA的蛋白水解作用的新兴生物学作用。此外，我们最近的组织蛋白酶L基因敲除小鼠的研究表明，这种蛋白酶在肾上腺中的NPY生产的参与。这些结果导致了本研究的下一阶段，其将（1）与PC 1和PC 2相比，评价分泌囊泡组织蛋白酶L对pro-NPY和CgA的体外和细胞加工，（2）评估组织蛋白酶L和PC酶在含有NPY和catestatin的分泌囊泡中的细胞和组织分布，（3）进行细胞反义和基因敲除研究以检查组织蛋白酶L和PC酶对于NPY和catestatin产生的相对作用，和（4）评价内桥蛋白2作为用于神经肽产生的组织蛋白酶L的内源性丝氨酸蛋白酶抑制剂。结果将证明组织蛋白酶L和内肽2的相对作用，相比PC 1和PC 2，在活性NPY和catestatin肽调节剂的生物合成。项目3补充了了解参与血压调节的交感神经效应物的调节这一项目主题。

Neuropeptide Y (NPY) and catestatin peptides are secreted from adrenomedullary chromaffin cells and sympathetic nerves for the regulation of blood pressure. NPY acts as a direct vasoconstrictor, and catestatin functions as an autocrine regulator to inhibit nicotine-stimulated catecholamine release. Elevated NPY and reduced catestatin in essential hypertension implicate their participation as neuroeffectors in regulating blood pressure. Importantly, knowledge of the major proteolytic enzymes (s) responsible for converting their respective prohormone precursors into active NPY and catestatin is crucial for understanding regulatory mechanisms that control blood pressure. The prohormone precursors of NPY and catestatin, pro-NPY and chromogranin A (CgA), respectively, undergo proteolytic processing within secretory vesicles of adrenal medulla, known as chromaffin granules. We have identified secretory vesicle cathepsin L, previously known as ?prohormone thiol protease? (PTP), as a key processing enzyme for pro-NPY and CgA. Moreover, this project discovered a novel endogenous serpin, endopin 2 that inhibits secretory vesicle cathepsin L. In addition, the subtilisin-like PC1 and PC2 proteases in secretory vesicles may also participate in pro-NPY and CgA processing. Based on these new findings, the goal of Project 3 will be to assess the roles of secretory vesicle cathepsin L and endopin 2, compared to PC1 and PC2, in the production of NPY and catestatin neuropeptides that regulate blood pressure. This project will test the hypothesis that secretory vesicle cathepsin L may be a major processing enzyme for NPY and catestatin, compared to PC1 and PC2 enzymes. Our new results support the emerging biological role of cathepsin L function in secretory vesicles for proteolysis of pro-NPY and CgA. Moreover, our recent studies of cathepsin L knockout mice suggest participation of this protease in NPY production in adrenals. These results lead to the next phase of this study that will (1) evaluate in vitro and cellular processing of pro-NPY and CgA by secretory vesicle cathepsin L, compared to PC1 and PC2, (2) assess the cellular and tissue distribution of cathepsin L and PC enzymes in secretory vesicles that contain NPY and catestatin, (3) conduct cellular antisense and gene knockout studies to examine the relative roles of cathepsin L and PC enzymes for NPY and catestatin production, and (4) evaluate endopin 2 as an endogenous serpin inhibitor of cathepsin L for neuropeptide production. Results will demonstrate the relative roles for cathepsin L and endopin 2, compared to PC1 and PC2, in the biosynthesis of active NPY and catestatin peptide regulators. Project 3 complements the program project theme of understanding the regulation of sympathetic neuroeffectors that participate in blood pressure regulation.