REGULATION OF THE PGH SYNTHASE GENES

PGH 合酶基因的调控

• 批准号: 2180544
• 负责人: David Lee DEWITT
• 金额: $ 18.07万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-07-01 至 1999-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2180544
• 关键词: antiinflammatory agents; biological models; clone cells; dexamethasone; eicosanoid metabolism; enzyme biosynthesis; enzyme structure; enzyme substrate complex; glucocorticoids; isozymes; laboratory rabbit; messenger RNA; posttranscriptional RNA processing; prostaglandin endoperoxide synthase; protein signal sequence; protein structure function; transfection

Prostaglandin endoperoxide H(PGH) synthase (cyclooxygenase) is a central enzyme in the prostaglandin biosynthetic pathway and also the primary site of action of aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs). Mammalian cells contain two related, but unique, forms of PGH synthase, referred to as PGH synthase-1 (PGHS-1) and PGH synthase-2 (PGHS02). These two enzymes appear to have specialized functions; PGHS-1 produces prostaglandins that act extracellularly s local hormones to coordinate short-term cellular responses to hormonal stimulation; while PGHS-2 produces prostaglandins that coordinate prolonged physiological events such as inflammation and ovulation. PGHS-2 produces prostaglandins that are directed into the nucleus where, we would predict, they also help regulate mitogenesis. The primary hypothesis that we propose to test in this application is that PGHS-1 and pGHS-2 form two separate prostaglandin biosynthetic pathways. Immunocytochemical localization by us indicates that PGHS-1 and PGHS-2 are located on different cellular membranes and produce prostaglandins that are shunted to different intracellular spaces. PGHS-1 is located predominantly in the endoplasmic reticulum and produces prostaglandins that are released into the cytoplasm where they presumably exit the cell to signal via extracellular receptors. PGHS-2 is located predominately on the nuclear membrane and releases prostaglandin on the nucleus and cytoplasm. How prostaglandin might signal in the nucleus is not known. Recently reported experiments indicate the PGHS-1 and PGHS-2 use separate arachidonate acid pools. To establish model systems with which to study the biochemical basis for segregation of the two prostaglandin synthase pathways, we are proposing to construct a set of 10 stably transfected cells which can inducibly express native and variously modified PGHS-1 and PGHS-2 proteins. These cell lines will be used to study the structure-function relationships that determine substate coupling, product channeling, and intracellular localization of PGHS-1 and PGHS-1 and PGHS-2 (Specific Aims 1-2), and also to examine the protein and mRNA sequences responsible for post-transcriptional regulation of PGHS-2 mRNA and protein stability (Specific Aims 3-4). If separate systems for prostaglandin synthesis do exist, with different mechanisms for arachidonate release and different effector pathways, these pathways may provide additional sites for therapeutic intervention of inflammation. in addition, a more complete understanding of the cellular roles for prostaglandin synthesis by PGHS-1 and PGHS-2 may help us to better understand the protective effects of NSAIDs on certain cancers.

前列腺素内过氧化物H（PGH）合酶（环氧合酶）是一种中枢性的 前列腺素生物合成途径中的酶，也是 阿司匹林和其他非甾体抗炎药的作用 （NSAID）。 哺乳动物细胞含有两种相关但独特的PGH形式 PGH合酶，称为PGH合酶-1（PGHS-1）和PGH合酶-2 （PGHS 02）。 这两种酶似乎有专门的功能; PGHS-1 产生胞外作用于局部激素的胡枝子素， 协调对激素刺激的短期细胞反应; PGHS-2产生与延长的生理 炎症和排卵等事件。 PGHS-2产生三尖杉酯碱 我们可以预测，它们也会帮助 调节有丝分裂。 我们要检验的主要假设是 该应用是PGHS-1和pGHS-2形成两个单独的前列腺素 生物合成途径 免疫细胞化学定位显示 PGHS-1和PGHS-2位于不同的细胞膜上， 产生被分流到不同的细胞内空间的木兰素。 PGHS-1主要位于内质网中并产生 它们被释放到细胞质中， 通过细胞外受体发出信号。 PGHS-2位于 主要在核膜上释放前列腺素， 细胞核和细胞质。 前列腺素如何在细胞核中发出信号 不知道。 最近报道的实验表明，PGHS-1和PGHS-2 使用分开的花生四烯酸酸池。 建立模型系统， 从而研究两者分离的生化基础 前列腺素合酶途径，我们建议构建一组10个 稳定转染的细胞，其可诱导表达天然的和多种 修饰的PGHS-1和PGHS-2蛋白。 这些细胞系将用于 研究决定子状态的结构-功能关系 PGHS-1的偶联、产物通道和细胞内定位， PGHS-1和PGHS-2（特异性目的1-2），并且还检查蛋白质和 负责PGHS-2转录后调节的mRNA序列 mRNA和蛋白质稳定性（特异性目的3-4）。 如果单独的系统 前列腺素的合成确实存在，不同的机制， 花生四烯酸释放和不同的效应途径，这些途径可能 为炎症的治疗性干预提供额外的位点。 在 此外，更全面地了解细胞的作用， PGHS-1和PGHS-2合成前列腺素可能有助于我们更好地 了解NSAID对某些癌症的保护作用。