REGULATION OF PROLACTIN GENE EXPRESSION

总的目标是理解分子事件所涉及的 催乳素基因的组织特异性的、受调控的表达。 催乳素基因在转录水平上受一种 激素的数量，包括雌二醇，表皮生长因子， 促甲状腺激素释放激素和多巴胺。建议进行的研究 寻求检查允许多激素的机制 催乳素转录的调控。特别关注 将集中在催乳素基因的细胞内调节上 转录，对催乳素基因重要的DNA序列 转录和与之相互作用的DNA结合因子 序列。具体目标和途径包括：1)角色定位 CAMP依赖的蛋白激酶催化亚单位在 介导cAMP对催乳素基因转录的影响将是 通过使用一种编码热稳定的合成基因进行检查 2)雌激素的结构特征 大鼠催乳素基因的反应元件 结合雌二醇受体并刺激转录 将通过构建合成雌激素进行检查 反应元件及反应元件的诱变作用 雌激素受体DNA结合域的特征 对与催乳素雌激素反应的相互作用很重要 将通过对克隆的受体进行突变来探索元素 CDNA.4)链接子扫描突变将用于解剖 特定DNA序列在紧邻5‘侧翼区中的作用 和催乳素的上游增强子/雌激素反应元件 基因5)与特定催乳素相互作用的DNA结合蛋白 基因序列将通过迁移率变化分析进行鉴定，纯化 通过亲和层析和克隆的cDNA。这些研究应该 加深对所涉及的一般机制的了解 在基因表达的调控以及特异性 调节这种生理上重要的荷尔蒙。

The overall goal is to understand the molecular events involved in the tissue-specific, regulated expression of the prolactin gene. The prolactin gene is regulated at the transcriptional level by a number of hormones including estradiol, epidermal growth factor, thyrotropin releasing hormone and dopamine. The proposed studied seek to examine the mechanisms which permit the multihormonal regulation of prolactin transcription. In particular attention will be focused on intracellular regulators of prolactin gene transcription, DNA sequences which are important for prolactin gene transcription and on DNA-binding factors which interact with these sequences. The specific aims are approaches include: 1) The role of the catalytic subunit of the cAMP-dependent protein kinase in mediating cAMP effects on prolactin gene transcription will be examined by using a synthetic gene coding for the heat stable inhibitor of the kinase 2) Structural features of the estrogen response element of the rat prolactin gene which are necessary for binding the estradiol receptor and for stimulation of transcription will be examined through construction of synthetic estrogen response elements and mutagenesis of the response element 3) Features of the DNA binding domain of the estrogen receptor which are important for interaction with the prolactin estrogen response element will be explored through mutagenesis of the cloned receptor cDNA. 4) Linker scanning mutagenesis will be used to dissect the role of specific DNA sequences in the immediate 5' flanking region and an upstream enhancer/estrogen response element of the prolactin gene 5) DNA binding proteins which interact with specific prolactin gene sequences will be identified by mobility shift assay, purified by affinity chromatography and cDNAs cloned. These studies should provide increased understanding of the general mechanisms involved in the regulation of gene expression as well as the specific regulation of this physiologically important hormone.