PAIRBP & PGRMC1 act as a membrane receptor complex to mediate P4's ovarian action

对BP

基本信息

批准号：
7924132
负责人：
JOHN J PELUSO
金额：
$ 30.82万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-09-01 至 2012-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Our previous studies have identified two proteins, referred to as PAIRBP1 and PGRMC1, that are expressed by granulosa and luteal cells. We propose that these two proteins interact to form a membrane complex that functions as a membrane receptor for progesterone (P4). We further propose that this P4 receptor complex mediates the non-genomic actions of P4. These actions include inhibiting mitosis, maintaining viability and regulating the steroidogenic capacity of granulosa and luteal cells. In this grant proposal we will present a series of experiments designed to address the following three specific aims: To determine the role of PAIRBP1 and PGRMC1 in regulating P4's biological actions. This will be done using siRNA and over expression to deplete and increase levels of these two proteins. The effect of changing the levels of these proteins on P4's ability to bind 3H-P4, inhibit mitosis and apoptosis and modulate steroid secretion will be monitored. These studies will be conducted on rat granulosa cells, rat luteal and human granulosa/luteal cells. To determine the function of the PAIRBP1 in the PAIRBP1-PGRMC1 complex. There are at least three possible mechanisms through which PAIRBP1 can interact with PGRMC1 to influence P4's actions. These include interacting with PGRMC1 to 1) form an "optimal binding pocket" for P4; 2) organize PGRMC1 and other unknown proteins into a large complex that is required for P4 signaling and 3) promote PGRMd's localization to the plasma membrane. To determine the molecular site required for the formation of the PAIRBP1-PGRMC1 complex and whether PAIRBP1-PGRMC1 interaction is required to mediate P4's action. Studies will be conducted to determine whether these two proteins bind to each other directly or indirectly via an intermediary protein. The amino acid sequence in PGRMC1 that is responsible for the formation of this complex will be identified. Specific strategies will also be developed to disrupt the PAIRBP1- PGRMC1 complex. These strategies will be based on the amino acid sequence within PGRMC1 that is involved in forming this complex. Then the effect of disrupting the PAIRBP1-PGRMC1 complex on P4's ability to regulate ovarian cell function will be assessed. If correct, this concept will change our understanding of P4's role in regulating ovarian function. It will also point the way toward the development on novel pharmacological agents that could selectively influence the PAIRBP1-PGRMC1 receptor complex and thereby block P4's non-genomic actions without altering the actions of the nuclear P4 receptor. These new pharmacological agents could have utility in the development contraceptives and in treating infertility and some forms of cancer.

描述（由申请人提供）：我们之前的研究已经鉴定出两种蛋白质，称为 PAIRBP1 和 PGRMC1，由颗粒细胞和黄体细胞表达。我们认为这两种蛋白质相互作用形成膜复合物，充当孕酮 (P4) 的膜受体。我们进一步提出该 P4 受体复合物介导 P4 的非基因组作用。这些作用包括抑制有丝分裂、维持活力和调节颗粒细胞和黄体细胞的类固醇生成能力。在本拨款提案中，我们将提出一系列实验，旨在解决以下三个具体目标：确定 PAIRBP1 和 PGRMC1 在调节 P4 生物作用中的作用。这将通过使用 siRNA 和过度表达来消除和增加这两种蛋白质的水平来完成。将监测改变这些蛋白质的水平对P4结合3H-P4、抑制有丝分裂和细胞凋亡以及调节类固醇分泌的能力的影响。这些研究将在大鼠颗粒细胞、大鼠黄体和人颗粒/黄体细胞上进行。确定 PAIRBP1-PGRMC1 复合物中 PAIRBP1 的功能。 PAIRBP1 可以通过至少三种可能的机制与 PGRMC1 相互作用来影响 P4 的行为。这些包括与 PGRMC1 相互作用，以 1) 形成 P4 的“最佳结合袋”； 2) 将 PGRMC1 和其他未知蛋白质组织成 P4 信号传导所需的大型复合物，3) 促进 PGRMd 定位到质膜。确定形成 PAIRBP1-PGRMC1 复合物所需的分子位点，以及介导 P4 的作用是否需要 PAIRBP1-PGRMC1 相互作用。将进行研究以确定这两种蛋白质是否直接或通过中间蛋白质间接相互结合。将鉴定 PGRMC1 中负责形成该复合物的氨基酸序列。还将制定具体策略来破坏 PAIRBP1-PGRMC1 复合体。这些策略将基于参与形成该复合物的 PGRMC1 内的氨基酸序列。然后将评估破坏 PAIRBP1-PGRMC1 复合物对 P4 调节卵巢细胞功能的能力的影响。如果正确的话，这个概念将改变我们对P4在调节卵巢功能中的作用的理解。它还将为开发新型药物制剂指明道路，这些药物可以选择性地影响 PAIRBP1-PGRMC1 受体复合物，从而阻断 P4 的非基因组作用，而不改变核 P4 受体的作用。这些新药理制剂可用于开发避孕药具以及治疗不孕症和某些形式的癌症。