REGULATION OF GENE EXPRESSION IN THE ADENOVIRUS SYSTEM AND IN TRANSGENIC MICE

腺病毒系统和转基因小鼠中基因表达的调控

• 批准号: 4693712
• 负责人: H WESTPHAL
• 金额: --
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/4693712
• 关键词: Adenoviridae; Retroviridae; cell transformation; developmental genetics; electron microscopy; embryo /fetus cell /tissue; eukaryote; gene expression; genetic manipulation; genetic mapping; genetic transcription; genetic translation; helper virus; messenger RNA; molecular cloning; molecular pathology; nucleic acid structure; oncogenic virus; plasmids; simian virus 40; tissue /cell culture; transforming virus; viral carcinogenesis; virus DNA; virus RNA; virus genetics; virus infection mechanism

Our laboratory investigates mechanisms of gene control in mammalian cells in culture and in the whole animal. One of two major projects, dealing with the E1A regulatory function of adenovirus, has been completed. The E1A function acts as a transcriptional modulator and is involved in malignant transformation. Sequences encoding E1A proteins or certain domains of these proteins have been inserted in prokaryotic expression vectors, and E1A proteins have been produced in E. coli. We microinjected the proteins into mammalian cells and measured their ability to migrate to the cell nucleus and to activate an adenovirus E1A deletion mutant. Information for nuclear localization and for viral gene activation has been shown to be encoded by distinct domains of the adenovirus E1A gene. Our second project involves insertion of specific gene constructs into mouse embryos. Spatial and temporal control of the expression of the inserted genes is examined in the resulting transgenic animals. So far, we have analyzed mice carrying three difference chimeric gene constructs. Each of these contains the gene for bacterial chloramphenicol acetyl transferase (CAT) under the control of a promoter/enhancer region derived from either the mouse Alpha2(I) collagen gene or from Rous sarcoma virus (RSV). The temporal and spatial contol of CAT expression in mice carrying the AlphaA crystallin-CAT or the Alpha2(I) collagen CAT construct reflects that of the genuine mouse genes from which the 5' flanking sequences of the chimeric genes were derived. In mice carrying the RSV-CAT construct CAT expression is preferentially directed to muscle and connective tissue. This reflects the disease specifically of sarcoma viruses. Finally, we have begun to analyze one RSV-CAT transgenic strain which is characterized by a dominant trait of embryonic lethality.

我们的实验室研究哺乳动物细胞基因控制的机制 在文化和整个动物中。 交易的两个主要项目之一 随着腺病毒的E1A调节功能，已经完成。 这 E1A功能充当转录调制器，参与 恶性转化。 编码E1A蛋白或某些的序列 这些蛋白质的结构域已插入核表达 载体和E1a蛋白已在大肠杆菌中产生。 我们微注射 蛋白质进入哺乳动物细胞，并测量其迁移到 细胞核并激活腺病毒E1A缺失突变体。 核定位和病毒基因激活的信息已有 被证明是由腺病毒E1A基因的不同域编码的。 我们的第二个项目涉及将特定基因构建体插入 小鼠胚胎。 表达的空间和时间控制 在产生的转基因动物中检查了插入的基因。 迄今为止， 我们已经分析了携带三个差异嵌合基因构建体的小鼠。 这些中的每一个都包含细菌氯霉素乙酰基的基因 在启动子/增强子区域的控制下转移酶（CAT） 从小鼠α2（i）胶原蛋白基因或ROUS肉瘤病毒中 （RSV）。 携带的小鼠猫表达的时间和空间抑制 Alphaa Crystallin-Cat或Alpha2（i）胶原蛋白CAT构建体反映 真正的小鼠基因的5'侧翼序列 嵌合基因得出。 在携带RSV-CAT构造的小鼠中 猫表达优先针对肌肉和结缔组织 组织。 这反映了特别是肉瘤病毒的疾病。 最后，我们开始分析一个RSV-CAT转基因菌株， 以胚胎致死性的主要特征为特征。