NOVEL CELL CYCLE REGULATION BY ADENOVIRUS E1B 55K

腺病毒 E1B 55K 对细胞周期的新型调控

• 批准号: 7909155
• 负责人: David Arnold Ornelles
• 金额: $ 13.56万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7909155
• 关键词: Accounting; Address; Adenovirus Infections; Adenoviruses; Attenuated; Cell Cycle; Cell Cycle Regulation; Cells; Clinical Trials; Complementary DNA; Defective Viruses; Employee Strikes; G1 Phase; Gene Expression; Genes; Goals; Growth; Human; Hybrid Cells; In Vitro; Infection; Malignant Neoplasms; Maps; Measures; Messenger RNA; Nature; Normal Cell; Oncolytic; Phase; Phase III Clinical Trials; Poly A; Protein Isoforms; Proteins; RNA Interference; RNA Splicing; RUNX1 gene; Recombinant DNA; Research; Research Personnel; Ribosomes; Rodent; Role; Scanning; Testing; Translating; Translations; Viral; Viral Genes; Viral Proteins; Virus; Virus Diseases; Work; base; human disease; killings; mutant; neoplastic cell; novel; overexpression; physical property; promoter; protein function; research study; response; tumor

DESCRIPTION (provided by applicant): Oncolytic ElB-55K-mutant adenoviruses have entered phase III clinical trials. However, the basis for their selective replication in tumor cells is poorly understood. Work of this lab has shown that ElB-55K-mutant viruses selectively replicate in and kill S phase cells better than G1 cells; this is likely to account for the selective replication in tumors. This research seeks to understand the tumor-selective nature of the E1B- 55K-mutant by understanding its S phase-selective and G1-restricted nature. Results from the previous period showed that late viral gene expression is suppressed in G1 cells at the level of translation, apparently in response to the E4orf1 or E4orf2 genes. The polysomal distribution of late viral mRNA in S phase and G1 cells infected with ElB-55K-mutant viruses suggests that late viral translation is restricted in G1 cells at the level of ribosome shunting. The E1B-55K protein of the wild-type virus promotes viral translation in association with the E4orl"6 protein and host cell factors. A candidate host cell factor has been mapped to the RUNX1 gene. Three specific aims are proposed in this application. Aim 1 seeks to determine how late viral translation is suppressed in G1-infected cells by comparing viral mRNA synthesis/ed in G1- and S phase-infected cells and by measuring the rates of key translational steps in synchronized and infected cells. This will test the hypothesis that ribosome shunting is decreased in the G1-infected cell. Aim 2 will elucidate the role of E4orfl and E4orf2 in attenuating late viral translation and restricting virus growth in G1. Viruses defective in either E4orlT or E4orf2 and the E1B-55K genes will be prepared and evaluated with respect to the cell cycle-restriction. Translation directed by ribosome scanning and ribosome shunting will he compared in cells infected with cell cycle-restricted and non-restricted mutant viruses to test the hypothesis that E4orf 1 attenuates translation at the level of ribosome shunting and that this is the basis for the G1-restriction. Aim 3 will identify the RUNX1 isoform important for E1B-55K function and determine if its activity during an infection is cell cycle-dependent. Candidate isoforms will be over expressed to determine if they compensate for the loss of E1B-55K function and RNA interference will be used to ablate RUNX1 expression to determine the importance of RUNX1 to adenovirus infection.

描述（由申请人提供）：溶瘤性elb - 55k突变腺病毒已进入III期临床试验。然而，它们在肿瘤细胞中选择性复制的基础尚不清楚。本实验室的工作表明，elb - 55k突变病毒选择性地在S期细胞中复制并杀死比G1期细胞更好；这可能是肿瘤中选择性复制的原因。本研究旨在通过了解E1B- 55k突变体的S期选择性和g1限制性来了解其肿瘤选择性。前期的研究结果表明，G1细胞在翻译水平上抑制了晚期病毒基因的表达，这显然是对E4orf1或E4orf2基因的反应。在感染elb - 55k突变病毒的S期和G1细胞中，晚期病毒mRNA的多体分布表明，晚期病毒翻译在G1细胞中受到核糖体分流水平的限制。野生型病毒的E1B-55K蛋白与E4orl ' 6蛋白和宿主细胞因子相关，促进病毒转译。候选宿主细胞因子已被定位到RUNX1基因。在此应用程序中提出了三个具体目标。Aim 1旨在通过比较G1期和S期感染细胞中的病毒mRNA合成/ed，以及测量同步和感染细胞中关键翻译步骤的速率，确定G1感染细胞中晚期病毒翻译是如何被抑制的。这将验证g1感染细胞中核糖体分流减少的假设。目的2将阐明E4orfl和E4orf2在G1中减弱病毒晚期转译和限制病毒生长中的作用。将制备E4orlT或E4orf2和E1B-55K基因缺陷的病毒，并对其细胞周期限制进行评估。通过核糖体扫描和核糖体分流引导的翻译将在受细胞周期限制和非限制突变病毒感染的细胞中进行比较，以检验E4orf 1在核糖体分流水平上减弱翻译的假设，这是g1限制的基础。目的3将鉴定对E1B-55K功能重要的RUNX1亚型，并确定其在感染期间的活性是否依赖于细胞周期。候选异构体将被过表达以确定它们是否补偿E1B-55K功能的丧失，RNA干扰将被用来去除RUNX1的表达以确定RUNX1对腺病毒感染的重要性。

项目成果

Serotype-specific reorganization of the Mre11 complex by adenoviral E4orf3 proteins.

腺病毒 E4orf3 蛋白对 Mre11 复合物进行血清型特异性重组。

• DOI: 10.1128/jvi.79.11.6664-6673.2005
• 发表时间: 2005
• 期刊: Journal of virology
• 影响因子: 5.4
• 作者: Stracker,TravisH;Lee,DarwinV;Carson,ChristianT;Araujo,FelipeD;Ornelles,DavidA;Weitzman,MatthewD
• 通讯作者: Weitzman,MatthewD

An activity associated with human chromosome 21 permits nuclear colocalization of the adenovirus E1B-55K and E4orf6 proteins and promotes viral late gene expression.

与人类 21 号染色体相关的活性允许腺病毒 E1B-55K 和 E4orf6 蛋白在核内共定位，并促进病毒晚期基因表达。

• DOI: 10.1128/jvi.77.14.8087-8098.2003
• 发表时间: 2003
• 期刊: Journal of virology
• 影响因子: 5.4
• 作者: Chastain-Moore,AmyM;Roberts,Terry;Trott,DeborahA;Newbold,RobertF;Ornelles,DavidA
• 通讯作者: Ornelles,DavidA

Diverse roles for E4orf3 at late times of infection revealed in an E1B 55-kilodalton protein mutant background.

E1B 55 千道尔顿蛋白突变体背景揭示了 E4orf3 在感染后期的不同作用。

• DOI: 10.1128/jvi.78.18.9924-9935.2004
• 发表时间: 2004
• 期刊: Journal of virology
• 影响因子: 5.4
• 作者: Shepard,RobinN;Ornelles,DavidA
• 通讯作者: Ornelles,DavidA