PITUITARY KALLIKREIN & PROHORMONE PROCESSING

垂体激肽释放酶

• 批准号: 3231131
• 负责人: CLAUD A POWERS
• 金额: $ 19.01万
• 依托单位: NEW YORK MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-04-01 至 1993-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3231131
• 关键词: affinity chromatography; bioassay; cell components; electron microscopy; enzyme induction /repression; enzyme mechanism; enzyme substrate; gel electrophoresis; high performance liquid chromatography; hormone regulation /control mechanism; immunochemistry; kallikreins; laboratory mouse; laboratory rabbit; laboratory rat; mammary gland; mitogens; peptidases; pituitary gland; prolactin; proopiomelanocortin; protein sequence; radioimmunoassay; urinalysis

Tissue (glandular) kallikrein (TK) is a trypsin-like serine protease which can generate biologically active peptides from inactive precursors with high specificity. I have recently discovered TK in the rat pituitary, and have found that TK exhibits patterns of tissue specific expression and physiological regulation closely paralleling MSH and beta-endorphin in the intermediate lobe, and prolactin (PRL) in the anterior pituitary lobe (AP). The proposed research will examine TK's role in the posttranslational processing of PRL in the AP, and Pro-opiomelanocortin (POMC; the precursor to MSH and beta-endorphin) in the intermediate lobe. Rat pituitary TK will be purified and its physiochemical and enzymatic properties compared to TK from other tissues. The ability of TK to cleave PRL and POMC will be examined using gel-electrophoresis and HPLC, and cleavage sites identified by microsequencing. PRL processing by TK may result in novel hormones with important functions in reproductive biology and mammary growth. Therefore, PRL cleavage products will be bioassayed for classic PRL actions (induction of milk-producing enzymes and stimulation of Nb2 lymphoma cell growth), as well as novel mitogenic actions on rat mammary glands in vivo. The cellular and subcellular localization of TK and cleaved PRL in the rat pituitary will be examined using immunohistochemistry and subcellular fractionation to see if TK is localized in sites of prohormone processing. Changes in TK, PRL and POMC levels, biosynthesis, and secretion will be compared following various physiological, hormonal and pharmacological in vivo and in vivo and in vitro to determine how tightly their regulation is interlinked. Alternative functions for rat pituitary TK will be tested (e.g., kinin generation and kinin modulation of pituitary hormone release). The results will determine whether TK functions to process peptides other than kinins, may elucidate an important mechanism of hormone biosynthesis, and reveal a novel role for PRL folding intermediates. Demonstration of an estrogen- induced pathway for generating PRL fragments with novel mitogenic activity on the mammary gland may be of considerable importance to current theories of the hormonal control of mammary growth and tumorigenesis in both rats and man.

组织（腺）激肽释放酶 (TK) 是一种胰蛋白酶样丝氨酸 蛋白酶可产生生物活性肽 具有高特异性的非活性前体。 我最近有 在大鼠垂体中发现了 TK，并发现 TK 表现出 组织特异性表达和生理调节模式 中间体中的 MSH 和 β-内啡肽密切相关 垂体前叶 (AP) 和催乳素 (PRL)。 这 拟议的研究将检验传统知识在翻译后的作用 AP 中 PRL 的加工，以及阿片黑皮质素原 (POMC； MSH 和 β-内啡肽的前体）位于中间叶。 鼠 垂体TK将被纯化，其理化和酶学 与其他组织的 TK 相比的特性。 TK的能力 将使用凝胶电泳检查切割 PRL 和 POMC 和 HPLC，以及通过微测序鉴定的切割位点。 催乳素 TK 加工可能会产生具有重要作用的新型激素 在生殖生物学和乳腺生长中发挥作用。 所以， PRL 裂解产品将进行经典 PRL 作用的生物测定 （诱导产奶酶和刺激 Nb2 淋巴瘤细胞生长），以及对大鼠的新的有丝分裂作用 体内的乳腺。 细胞和亚细胞定位 将使用以下方法检查大鼠垂体中的 TK 和裂解 PRL 免疫组织化学和亚细胞分级分离以确定 TK 是否有效 位于激素原加工位点。 TK、PRL 的变化 和 POMC 水平、生物合成和分泌将进行比较 遵循各种生理、激素和药理作用 体内和体外以确定它们的紧密程度 监管是相互关联的。 大鼠垂体的替代功能 将测试 TK（例如，激肽生成和激肽调节） 垂体激素释放）。结果将决定是否TK 处理除激肽以外的肽的功能，可以阐明 激素生物合成的重要机制，并揭示了一种新的 PRL 折叠中间体的作用。 雌激素的示范- 产生具有新型有丝分裂作用的PRL片段的诱导途径 对乳腺的活动可能相当重要 目前关于乳腺生长的激素控制的理论 大鼠和人的肿瘤发生。