RETROVIRUS REGULATION IN VITRO AND DURING DEVELOPMENT

体外和开发过程中的逆转录病毒监管

• 批准号: 2180916
• 负责人: KATHLEEN F CONKLIN
• 金额: $ 18.69万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-07-01 至 1996-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2180916
• 关键词: Retroviridae; Retroviridae disease; avian leukosis virus; binding proteins; chick embryo; gene expression; genetic enhancer element; genetic promoter element; genetic regulation; genetic transcription; in situ hybridization; infection related neoplasm /cancer; microorganism genetics; neoplastic transformation; northern blottings; nucleic acid repetitive sequence; nucleic acid sequence; provirus; regulatory gene; tissue /cell culture; transcription factor; virus DNA; virus assembly; virus replication

The goal of experiments in this proposal is to determine the structural and functional relationship between the avian leukosis-sarcoma virus enhancer and enhancer sequences within the LTRs of avian endogenous viruses (evs). Non-acute transforming avian leukosis viruses (ALVs) induce a high incidence of bursal lymphomas after infection of certain susceptible strains of chickens. Cellular transformation by these viruses is commonly associated with integration of proviral DNA near or within the cellular oncogene c-myc which results in increased expression of this gene under the control of promoter/enhancer sequences within the U3 region of the proviral LTR. In contrast, the avian endogenous viruses (evs) are weakly oncogenic in vivo. It has been suggested that low level oncogenicity of avian evs is due primarily to the absence of a strong enhancer in their LTRs that prevents them from efficiently mediating cis-activation events such as those seen in exogenous virus-induced tumors. Unexpectedly however, sequences have been identified in ev LTRs that are able to replace, in an orientation independent mariner, essential enhancer sequences in the exogenous virus LTR with which they share limited sequence similarity. By constructing ev-exogenous virus hybrid LTRs, the sequences within ev LTRs required for this effect are being localized. Preliminary data has also been obtained indicating that ev enhancer sequences interact with cellular proteins distinct from exogenous virus enhancer binding proteins. Thus, enhancer motifs within the ev LTR appear distinct from those in the exogenous virus LTR. The availability of hybrid LTRs that are transcribed efficiently yet contain enhancer motifs from the weakly oncogenic ev provides the opportunity to directly test the relationships between enhancer activity, the identity and function of distinct enhancer motifs, and oncogenesis. This will be investigated using a combined approach of protein binding studies and functional analyses on individual ev and exogenous virus enhancer domains and on ev-exogenous virus hybrid LTRs. Selected hybrid LTRs will then be used to generate replication competent virus in order to determine their growth rates in vitro and to generate virus stocks for future use in oncogenicity testing. The long range goal of these studies is to define and to potentially distinguish between regulatory elements that mediate enhancer activity in in vitro experiments and, in the future, those that are required for high level oncogenesis in vivo.

本提案中的实验目标是确定结构 禽白血病肉瘤病毒与禽流感病毒之间的关系 增强子和增强子序列内的禽内源性 病毒（EVs）。 非急性转化型禽白血病病毒（ALV） 在感染某些病毒后， 易感鸡种 细胞转化通过这些 病毒通常与前病毒DNA的整合相关， 在细胞癌基因c-myc中， 该基因的启动子/增强子序列的控制下， 前病毒LTR的U3区。 与此相反， (evs)在体内是弱致癌的。 有人说，低水平 禽EV的致癌性是由于 主要是因为它们的LTR中缺乏一个强大的增强子， 他们从有效地介导顺式激活事件，如那些看到 外源性病毒诱导的肿瘤。 然而，出乎意料的是，已经在ev LTR中鉴定出序列， 能够取代，在一个方向独立的水手，必不可少的 它们与外源病毒LTR中的增强子序列共享 有限的序列相似性。 通过构建ev-外源病毒杂交体 LTR，该效应所需的ev LTR内的序列被 本地化。 初步数据也表明， 增强子序列与细胞蛋白质相互作用，不同于 外源病毒增强子结合蛋白。因此， ev LTR似乎与外源病毒LTR中的不同。 的 有效转录但含有 来自弱致癌EV的增强子基序提供了机会， 直接测试增强子活性、同一性 和不同增强子基序的功能以及肿瘤发生。 这将是 使用蛋白结合研究的组合方法进行研究， 单个ev和外源病毒增强子结构域的功能分析 和在EV-外源病毒杂合LTR上。 选定的混合LTR将 用于产生有复制能力的病毒，以确定它们的 体外生长率，并产生病毒储备，供将来用于 致瘤性试验。 这些研究的长期目标是确定 并潜在地区分调节元件， 在体外实验中增强活性，在未来，那些 是体内高水平肿瘤发生所必需的。