REGULATION OF SIMIAN FOAMY VIRUS GENE EXPRESSION

猿泡沫病毒基因表达的调控

• 批准号: 3145532
• 负责人: Paul Luciw
• 金额: $ 17.66万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-07-01 至 1997-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3145532
• 关键词: CHO cells; Retroviridae; antibody; gene expression; genetic manipulation; genetic regulation; molecular cloning; nucleic acid repetitive sequence; plasmids; posttranscriptional RNA processing; transcription factor; virion; virus protein

The foamy viruses, or syncytium-forming viruses, are members of the spumaviriniae sub-family of retroviruses. Foamy viruses are found in many mammalian species and appear to be non-pathogenic in their natural hosts even though they have a wide tissue range and induce extensive cytopathology in cell cultures. The genomes of simian foamy virus type 1 (SFV-l) from rhesus macaque and human spumaretrovirus (HSRV) have been molecularly cloned and sequenced; pairwise comparisons of these two viruses show from 30% to 80% homology, depending on the region selected for analysis. Both viruses encode several open reading frames (ORFs) in addition to genes for virion structural proteins. Recent studies have revealed that SFV-1 and HSRV each encode a transactivator (taf) which functions to strongly augment transcription directed by the viral long terminal repeat (LTR). The main objective of this proposal is to elucidate molecular mechanisms regulating SFV-1 gene expression directed by the viral LTR. It remains to be determined whether taf acts directly on the LTR or whether transactivation is mediated by cellular factors. The target (TAR) for taf is distributed over two parts of the U3 domain of the viral LTR upstream from the TATA box in the viral promoter. Each these two TAR regions is about 300 base pairs (bp) long and no homologies (i.e., repeat elements) are detected between the two TARs. In addition, the promoter-distal TAR (TAR-d) functions in only in the sense orientation whereas the promoter-proximal TAR (TAR-p) functions in either orientation. These observations (on the complex nature of the targets) support the hypothesis that taf may function through more than one cellular factor which interacts with different sequence elements in the U3 region of the LTR. SPECIFIC AIM 1: Binding sites for cellular factors and the precise target sequences for taf in the SFV-1 LTR will be defined. SPECIFIC AIM 2: Monospecific antibodies will be made to detect the SFV-1 taf gene product; taf protein in infected cells will be extensively characterized. SPECIFIC AIM 3: Genetically engineered yeast and mammalian cells will be used to produce the SFV-1 taf gene product for structural and functional studies. SPECIFIC AIM 4: To elucidate the mechanism of SFV-1 transcriptional transactivation, functional domains of taf will be identified by mutagenesis and by evaluation of hybrid transactivators. These studies on SFV-1 regulation are relevant for elucidating mechanisms which control viral latency (or persistent infection) in the host animal. Observations made in the course of the proposed research will also enhance the understanding of mechanisms controlling eucaryotic transcription; the proposed studies on SFV-1 gene expression are significant since new (cellular) transcriptional factors may be identified.

泡沫病毒，或合胞体形成病毒，是 海绵病毒亚科逆转录病毒。泡沫状病毒在 许多哺乳动物物种，似乎在他们的天然非致病 寄主，尽管它们具有广泛的组织范围并诱导广泛的 细胞培养中的细胞病理学。猴泡沫病毒1型基因组的研究 来自恒河猴的L病毒和人类Spumaretrov已经被 分子克隆和测序；这两者的配对比较 病毒的同源性在30%到80%之间，具体取决于所选择的区域 以供分析。这两种病毒都编码了几个开放阅读框架(ORF) 病毒粒子结构蛋白基因的补充。最近的研究表明 揭示了SFV-1和HSRV各自编码一个反式激活因子(Taf)，Taf 强烈增强病毒Long引导的转录的功能 终端重复(LTR)。这项建议的主要目的是 定向调控SFV-1基因表达的分子机制 被病毒LTR感染。Taf是否直接起作用还有待确定 或者反式激活是否由细胞因子介导。 Taf的目标(TAR)分布在U3域的两个部分 病毒启动子中TATA盒上游的病毒LTR。每个人 这两个TAR区长约300个碱基对，没有同源性 (即，在两个TAR之间检测到重复元件)。此外, 启动子-远端TAR(tar-d)仅在某种意义上起作用 而启动子-近端TAR(TAR-P)在以下两种情况下都起作用 定位。这些观察(关于目标的复杂性质) 支持Taf可能通过不止一个途径发挥作用的假设 与不同序列元件相互作用的细胞因子 LTR的U3区。 特异性目标1：细胞因子的结合位点和精确靶点 将定义SFV-1 LTR中Taf的序列。 特异性目的2：制备单特异性抗体用于检测SFV-1 Taf基因产物；Taf蛋白在感染细胞中将广泛表达 特色化的。 特定目的3：基因工程酵母和哺乳动物细胞将 用于生产SFV-1 Taf基因产物，用于结构和功能 学习。 特异性目的4：阐明SFV-1转录调控机制 Taf的反式激活、功能结构域将通过 突变和杂合反式激活剂的评价。 这些关于SFV-1调控的研究对于阐明机制是相关的。 它们控制宿主动物中的病毒潜伏期(或持续感染)。 在拟议的研究过程中提出的意见也将 加深对真核生物调控机制的理解 转录；建议对SFV-1基因表达的研究包括 由于新的(细胞)转录因子可能是 确认身份。