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

对BP

基本信息

批准号：
8134344
负责人：
JOHN J PELUSO
金额：
$ 29.76万
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 2013-08-31
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）：我们以前的研究已经鉴定出了两种蛋白质，称为PairBP1和PGRMC1，这些蛋白质由颗粒和黄体细胞表达。我们建议这两种蛋白质相互作用以形成一种膜复合物，该膜复合物充当孕酮的膜受体（P4）。我们进一步提出，该P4受体复合物介导了P4的非基因组作用。这些作用包括抑制有丝分裂，维持生存力并调节颗粒和黄体细胞的类固醇生成能力。在此赠款建议中，我们将提出一系列旨在解决以下三个特定目的的实验：确定PARMBP1和PGRMC1在调节P4生物学作用中的作用。这将使用siRNA和过度表达来耗尽并增加这两种蛋白质的水平。将监测这些蛋白质水平对P4结合3H-P4，抑制有丝分裂和凋亡的能力的影响。这些研究将对大鼠颗粒细胞，大鼠黄体和人颗粒/黄体细胞进行。确定PAREBP1-PGRMC1复合物中PAIMBP1的功能。 PairBP1至少有三种可能的机制可以与PGRMC1相互作用以影响P4的作用。这些包括与PGRMC1与1）相互作用，形成了P4的“最佳结合口袋”； 2）将PGRMC1和其他未知蛋白组织成P4信号所需的大型复合物和3）促进PGRMD对质膜的定位。为了确定形成PARBBP1-PGRMC1复合物所需的分子位点，以及是否需要PARBBP1-PGRMC1相互作用来介导P4的作用。将进行研究以确定这两种蛋白是否通过中间蛋白直接或间接地相互结合。将鉴定出负责该复合物形成的PGRMC1中的氨基酸序列。还将制定特定的策略来破坏PAPBP1-PGRMC1复合物。这些策略将基于与形成这种复合物有关的PGRMC1中的氨基酸序列。然后，将评估破坏PARBP1-PGRMC1复合物对P4调节卵巢细胞功能能力的影响。如果正确，此概念将改变我们对P4在调节卵巢功能中的作用的理解。它还将指向新型药理学剂的发展，这些药理学剂可以选择性地影响PARMBP1-PGRMC1受体复合物，从而阻止P4的非基因组作用而不会改变核P4受体的作用。这些新的药理学剂可能在开发避孕药和不孕症和某些形式的癌症方面具有效用。