Regulation of Nox Enzymes by Calcium and Novel Subunits

描述(申请人提供)：我们最近描述了一个NADP家族 H-氧化酶，一种与癌症，特别是结肠癌和前列腺癌有关的NOx酶。这些酶是产生活性氧簇(ROS)的黄色素，其中一种是过氧化氢，具有信号分子的功能，刺激有丝分裂和血管生成。因此，这些酶在上皮细胞中的异常表达或激活被认为在癌症进展中发挥了作用。然而，到目前为止，人们对这些酶的活性是如何调节的知之甚少。因此，本应用重点研究了NOX1、NOX3、NOX4和NOX5的分子调控机制。NOx酶是吞噬细胞呼吸爆发氧化酶的催化亚基gp91Phox的同源物。后者受催化亚基p47Phox和p67Phox的调控，我们最近从结肠和睾丸的cDNA文库中发现了这两个亚基的新的同源物NOXO1(NOX组织蛋白1)和NOXA1(NOX激活蛋白1)。有证据表明，这些蛋白质是Nox1和Nox5的调节亚单位。我们鉴定了NOXO1的4种异构体(剪接体)和NOXAI的5种剪接体。此外，NOXO1和NOXA1含有Src同源3(SH3)结构域和富含脯氨酸的区域，NOXA1包含一个TPR基序的4个串联副本，该基序被认为参与与RAC或另一个小的GTP结合蛋白形成异二聚体。因此，这些亚基及其相互作用结构域在NoXL-5的调控组装/激活中的作用以及小GTP酶的作用将被研究。Nox5含有一个类似于钙结合蛋白Recovery in基团的结构域，推测受钙的调节。此外，Nox5含有一个富含脯氨酸的基序，据预测该基序与NOXO1中的SH3结构域相互作用。调节亚基和钙作为调控NOx5的替代或协同机制的作用将被研究。这些研究有望对与细胞生长和癌症相关的ROS信号的正常和异常产生产生影响。

DESCRIPTION (provided by applicant): We recently described a family of NADP H-oxidases, the Nox enzymes that have been implicated in cancer, particularly colon and prostate cancers. The enzymes are flavocytochromes that produce reactive oxygen species (ROS) and one of these, hydrogen peroxide, has functions as a signal molecule, stimulating mitosis and angiogenesis. Aberrant epithelial expression or activation of these enzymes has therefore been proposed to play a role in cancer progression. However, to date, little is known about how the activity of these enzymes is regulated. This application therefore focuses on the molecular mechanisms of regulation of Nox1, Nox3, Nox4 and Nox5. The Nox enzymes are homologs of gp91phox, the catalytic subunit of the phagocyte respiratory burst oxidase. The latter is regulated by catalytic subunits p47phox and p67phox, and we recently identified novel homologs of these subunits termed NOXO1 (Nox Organizing Protein 1) and NOXA1 (Nox Activating Protein 1) from colon and testis cDNA libraries. Evidence points to these proteins as regulatory subunits of the Noxl and possibly Nox5. We have identified 4 isoforms (splice forms) of NOXO1 and 5 splice forms of NOXAI. In addition, NOXO1 and NOXA1 contain Src-homology 3 (SH3) domains and proline-rich regions, and NOXA1 contains a 4 tandem copies of a TPR motif that is proposed to participate in heterodimer formation with Rac or another small GTP-binding protein. Thus, the role of these subunits and their interaction domains in the regulated assembly/activation of Noxl-5, and the role of small GTPases will be investigated. Nox5 contains a domain similar to the recoverin group of calcium-binding proteins, and is presumed to be regulated by calcium. In addition, Nox5 contains a proline-rich motif that is predicted to interact with an SH3 domain like the one in NOXO1. The role of regulatory subunits and calcium as alternative or synergistic mechanisms for regulating Nox5 will be investigated. These studies are expected to have implications with regard to both normal and aberrant generation of ROS signals relevant to cell growth and cancer.