FSH REGULATION OF A-KINASE ANCHORING PROTEINS

CAMP是第二个信使，它调节对许多不同 荷尔蒙和神经递质。CAMP水平升高导致 激活cAMP依赖的蛋白激酶(PKA)，从而产生 各种不同的生理反应。一个假说可以解释为什么 一种可以执行许多不同功能的酶是亚细胞 在每种不同的细胞类型中，PKA的定位受到严格控制。 因此，PKA将成为其目标或锚定在其附近 生理底物。PKA由四个亚单位组成，两个监管和 两种催化剂。PKA的锚定被认为是通过 蛋白激酶A调节亚单位(R)与特定蛋白的相互作用 细胞蛋白，称为A-激酶锚定蛋白(AKAPs)。 虽然PKA之间的蛋白质相互作用的特征 而锚定蛋白已经广泛存在，其生理功能 大多数AKAP仍然不为人知。颗粒细胞提供了一种理想的系统 研究AKAP的功能作用。众所周知，促卵泡生成素和 促黄体生成素促进颗粒细胞分化的机制 依赖于cAMP的产生和随后的激活 PKA.然而，PKA调节这种生物的机制 这一过程仍然难以捉摸。两个AKAP最近在 表达差异表达的原代颗粒细胞培养 受卵泡刺激素(FSH)调节。一的表达 AKAP 80蛋白是由FSH刺激的，而另一种蛋白的诱导 AKAP 140蛋白被FSH阻断。卵泡刺激素也会导致一种 蛋白激酶A亚基调节亚基(RIIbeta)的异构体及其转位 另一种PKA亚型(RIIpha)。所有这些数据都与 假设FSH促进PKA的再分布，可能是为了 细胞对第二种激素做出反应。Lh，这也会导致 营地的增加。为了检验这一假设，我们提出：L)分离和 表征AKAP 80和140；2)确定亚细胞 AKAP和RIIpha和RIIbeta的位置；以及3)确定 AKAPs 80和140及其亚型在颗粒细胞中的作用 要么通过阻断PKA与锚定蛋白的相互作用 使用锚定抑制肽或阻断它们的诱导 通过使用反义寡核苷酸获得蛋白质。这些研究将 提供对特定AKAP的生理作用的见解，以及新的 卵泡刺激素和促黄体生成素利用的细胞信号通路的信息 颗粒细胞。

cAMP is a second messenger which regulates responses to many different hormones and neurotransmitters. Elevated levels of cAMP results in the activation of the cAMP-dependent protein kinase (PKA) which produces a variety of distinct physiological responses. One hypothesis to explain how one enzyme can perform many different functions is that the subcellular localization of PKA is tightly controlled in each different cell type. Accordingly, PKA would be targeted or anchored adjacent to its physiological substrate. PKA consists of four subunits, two regulatory and two catalytic. The anchoring of PKA is believed to be accomplished through the interaction of the regulatory subunit (R) of PKA with specific cellular proteins, referred to as A-Kinase Anchoring Proteins (AKAPs). Although characterization of the protein-protein interaction between PKA and anchoring proteins has been extensive, the physiological function of most AKAPs remains unknown. Granulosa cells provide an ideal system for studying the functional role of AKAPs. It is well recognized that FSH and LH sequentially promote granulosa cell differentiation via a mechanism dependent upon the generation of cAMP and the subsequent activation of PKA. However, the mechanism by which PKA regulates this biological process remains elusive. Two AKAPs have recently been identified in primary granulosa cell cultures whose expression is differentially regulated by follicle stimulating hormone (FSH). The expression of one protein, AKAP 80, is stimulated by FSH, while the induction of the other protein, AKAP 140, is blocked by FSH. FSH also causes the induction of one isoform of PKA, regulatory subunit (RIIbeta) and the translocation of another isoform of PKA (RIIalpha). All of this data is consistent with the hypothesis that FSH promotes a redistribution of PKA, possibly to enable the cells to respond to a second hormone. LH, which also causes an increase in cAMP. To test this hypothesis we propose: l) to isolate and characterize both AKAP 80 and 140; 2) to determine the subcellular location of the AKAPs and RIIalpha and RIIbeta; and 3) to determine the functions of AKAPs 80 and 140 and isoforms of PKA in granulosa cells by either blocking the interaction of PKA with anchoring proteins through the use of anchoring inhibitor peptides or blocking the induction of these proteins through the use of antisense oligonucleotides. These studies will provide insights into the physiologic role of specific AKAPs and also new information on the cellular signaling pathway utilized by FSH and LH in granulosa cells.