FSH REGULATION OF A-KINASE ANCHORING PROTEINS

cAMP是第二信使，调节许多不同的反应。 激素和神经递质。cAMP水平升高导致 cAMP依赖性蛋白激酶（PKA）的激活， 各种不同的生理反应。有一个假说可以解释 一种酶可以执行许多不同的功能， PKA的定位在每种不同的细胞类型中受到严格控制。 因此，PKA将被靶向或锚定在其邻近区域。 生理底物PKA由四个亚基组成，两个调节亚基， 二是催化剂。PKA的锚定被认为是通过 PKA的调节亚基（R）与特异性 细胞蛋白，称为A-激酶转运蛋白（AKAP）。 虽然PKA与蛋白质间相互作用的特征 和锚定蛋白的生理功能已经广泛， 大多数AKAP仍然未知。颗粒细胞提供了一个理想的系统， 研究AKAP的功能作用。 众所周知，FSH和 LH通过一种机制依次促进颗粒细胞分化 依赖于cAMP的产生和随后的 PKA。 然而，PKA调节这种生物学效应的机制 进程仍然难以捉摸。最近确定了两个AKAP， 表达差异的原代颗粒细胞培养物 受促卵泡激素（FSH）调节。表达一 蛋白质AKAP 80被FSH刺激，而另一种蛋白质的诱导被FSH刺激。 AKAP 140蛋白被FSH阻断。FSH也会诱导一种 PKA亚型、调节亚基（RII β）和 PKA的另一种亚型（RII α）。所有这些数据都与 假设FSH促进PKA的重新分布，可能使 细胞对第二种激素产生反应。LH也会导致 cAMP增加。为了验证这一假设，我们提出：l）分离和 表征AKAP 80和140两者; 2）以确定亚细胞 AKAP和RIIalpha和RIIbeta的位置;以及3）确定 AKAP 80和140以及PKA亚型在颗粒细胞中的功能， 或者阻断PKA与锚定蛋白的相互作用， 锚定抑制剂肽或阻断这些肽的诱导的用途 蛋白质通过使用反义寡核苷酸。这些研究将 提供了对特定AKAP的生理作用的见解， 关于FSH和LH利用的细胞信号传导途径的信息， 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.