TRANSCRIPTIONAL REGULATION BY CAMP AND RETINOIDS

CAMP 和类维生素A 的转录调控

• 批准号: 2204251
• 负责人: Jodi Huggenvik
• 金额: $ 9万
• 依托单位: SOUTHERN ILLINOIS UNIVERSITY CARBONDALE
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-08-01 至 1998-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2204251
• 关键词: HeLa cells; autoradiography; gel mobility shift assay; genetic promoter element; genetic regulatory element; genetic transcription; molecular cloning; phosphopeptides; phosphorylation; protein isoforms; protein kinase A; receptor binding; recombinant proteins; reporter genes; retinoate; retinoid binding proteins; second messengers; site directed mutagenesis; tissue /cell culture; transcription factor; transfection

Retinoids are natural and synthetic derivatives of vitamin A (retinol) and have been recognized as physiologically important in cellular differentiation and proliferation. Retinoids are used extensively in the clinical treatment of a variety of dermatologic diseases and as chemotherapeutic anticancer agents. However, therapeutic doses can be teratogenic and cause embryonic death and fetal malformations in both humans and animals. Recent evidence suggested that the molecular mechanisms by which retinoids exert their teratogenic potential and physiological roles is through binding to nuclear retinoid receptors and altering gene expression. The retinoid receptors show structural homology and molecular mechanisms of action similar to the steroid-thyroid-vitamin D receptor superfamily. We have used the transient transfection of mammalian cells with a retinoic acid responsive reporter plasmid to study these mechanisms. The cotransfection of the plasmid expressing the catalytic subunit of cAMP-dependent protein kinase (PKA) revealed that the cAMP second messenger pathway of signal transduction is tightly coupled to the retinoid responsive pathway. We propose that retinoid receptors are natural substrates of PKA and this phosphorylation contributes to changes in gene expression. This hypothesis will be tested by: (i) confirmation of the phosphoamino acids in the RARs, (ii) in vitro mutagenesis to evaluate the effect of RAR phosphorylation on transcriptional activation, and (iii) the identification of other transcription factors, such as retinoid X receptor, that are regulated by PKA phosphorylation. These experiments should provide a better understanding of how two signalling pathways can elicit different responses depending on the participating transcriptional components in a cell. These observations may be applicable to a broad range of membrane receptors which are coupled to a second messenger system which can then interact with members of the intracellular steroid hormone receptor superfamily. It is expected that the results of our studies should provide the biomedical community with insights for better combined drug therapies.

类视网膜类似是维生素A（视黄醇）的天然和合成衍生物 在细胞中被认为在生理上很重要 分化和增殖。类维生素类似于广泛的 各种皮肤病疾病的临床治疗 化学治疗抗癌剂。但是，治疗剂量可以是 致死性并在两者中引起胚胎死亡和胎儿畸形 人类和动物。最近的证据表明分子 类视黄素发挥其致病潜力和的机制 生理作用是通过与核性类维生素类动物受体结合和 改变基因表达。 类维生素性受体显示结构同源性 和分子作用机制类似于类固醇 - 甲状腺维生素 D受体超家族。我们已经使用了 哺乳动物细胞具有视黄酸反应者质粒的哺乳动物细胞 这些机制。表达质粒的共转染 CAMP依赖性蛋白激酶（PKA）的催化亚基表明 训练营第二信使通路的信号转导通道紧密耦合 类维生素类动物的响应途径。 我们提出类视黄素受体是 PKA的天然底物和这种磷酸化有助于变化 在基因表达中。该假设将通过：（i）确认 RARS中的磷酸氨基酸，（ii）体外诱变以评估 RAR磷酸化对转录激活的影响，（III） 鉴定其他转录因子，例如类维生素类动物X 受体，受PKA磷酸化调节。这些实验 应该更好地了解两个信号通路如何 根据参与的转录引起不同的回答 细胞中的成分。这些观察结果可能适用于广泛的 耦合到第二使者系统的膜受体范围 然后可以与细胞内类固醇激素的成员相互作用 受体超家族。预计我们的研究结果 应该为生物医学界提供见解，以提供更好的结合 药物疗法。