STRUCTURAL AND FUNCTIONAL STUDIES OF MAMMALIAN ENDOGENOUS RETROVIRAL SEQUENCES

哺乳动物内源逆转录病毒序列的结构和功能研究

• 批准号: 3818206
• 负责人: A S KHAN
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 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/3818206
• 关键词: DNA binding protein; Retroviridae disease; aging; animal age group; chemical binding; gel electrophoresis; gene expression; genetic manipulation; genetic mapping; genetic regulation; genetic strain; genetic transcription; human age group; human subject; laboratory mouse; molecular cloning; murine leukemia virus; nucleic acid probes; nucleic acid sequence; transposon /insertion element; virus DNA; virus RNA; virus envelope; virus genetics; virus protein

Mammalian genomes contain multiple copies of retroviral-related sequences. The majority of these are defective for virus expression; however, some can produce subgenomic RNAs. Age- dependent expression of endogenous retroviral sequences was studied in Mus musculus wild mouse and man using endogenous MuLV and human 4-1 DNA probes. In both cases transcripts were detected in kidney, brain, and liver tissues, but expression was highest in the kidney. MuLV-related RNAs, 8.4 kb, 3.0 kb, and 1.8 kb were seen throughout life (18 day old embryo - 28 months of age) and increased with age. Transcripts, 3.0 kb and 1.8 kb, were seen in human tissues from 8 month fetus -85 years of age. These RNAs also appeared to increase with age. The results suggest a relaxation of gene control in mammalian cells with aging. A novel 5.0 kb retrotransponson-like element, B-13, was cloned from BALB/c mouse embryo DNA and sequenced. It was distantly related to known MuLV DNAs: <75% homology was seen in the gag and pol regions; env sequences were absent. Two other cloned DNAs isolated from the BALB/c mouse genome contained B-13-like genes but different flanking cellular sequences. Several copies of B-13- related genes were detected in inbred and closely related wild mouse genomes. These results indicate that B-13-like elements are amplified and dispersed in the mouse genome. Two novel protein binding sites were identified at the 5' end of all MuLV-related LTRs using 30 bp oligomers as probes in gel mobility shift assays. These highly conserved sites were present upstream of the enhancer in a region designated as the upstream conserved region (UCR) and bound nuclear factors designated as UCRF-L and UCRF-U. UCRF-L was the prominent species in murine tissue whereas UCRF-U predominated in cell lines. Northern analysis of 5 different mouse strains indicated a good inverse correlation between MuLV RNA expression and UCRF-L activity. This relationship was seen in case of both spontaneous and LPS-induced MuLV RNAs. These results suggest a negative regulatory function of UCRF-L in MuLV gene expression.

哺乳动物基因组含有多个拷贝的逆转录病毒相关的 序列的 其中大多数是有缺陷的病毒 然而，有些可以产生亚基因组RNA。 年龄- 研究了内源性逆转录病毒序列的依赖性表达 使用内源性MuLV和人在小家鼠野生小鼠和人中 4-1 DNA探针。 在这两种情况下，在肾脏中都检测到转录本， 脑和肝组织，但表达最高的是在肾脏。 MuLV相关RNA，8.4 kb，3.0 kb和1.8 kb，在整个细胞中可见 寿命（18天胚胎- 28个月），并随年龄增加。 在8例正常人的组织中分别检测到3.0kb和1.8kb的转录本， 月胎儿-85岁。 这些RNA似乎也增加了 随年龄 研究结果表明，基因控制的放松， 哺乳动物细胞的衰老。 从大肠杆菌中克隆了一个新的5.0 kb的逆转录转座子样元件B-13， BALB/c小鼠胚胎DNA并测序。 它是远亲 与已知的MuLV DNA：在gag和pol中观察到<75%的同源性 区域;不存在env序列。 分离出另外两个克隆DNA 来自BALB/c小鼠基因组的DNA含有B-13样基因， 不同的侧翼细胞序列 几份B-13- 在近交系和近缘野生系中检测到相关基因 老鼠的基因组 这些结果表明，B-13类元素是 扩增并分散在小鼠基因组中。 两个新的蛋白质结合位点被鉴定在5'端的 使用30 bp寡聚体作为凝胶中探针的所有MuLV相关LTR 迁移率变化测定。 这些高度保守的位点 在指定为上游增强子的区域中的增强子上游， 保守区（UCR）和结合的核因子， UCRF-L和UCRF-U。 UCRF-L是小鼠中的主要种属 而UCRF-U在细胞系中占主导地位。 北方 对5种不同小鼠品系的分析表明， MuLV RNA表达与UCRF-L活性之间的相关性。 这 在自发和LPS诱导的情况下均观察到相关性 MuLV RNA。 这些结果表明负调节功能 UCRF-L在MuLV基因表达中的作用。