Walleye Dermal Sarcooma Virus Accessory Protein Function

白斑眼真皮肉瘤病毒辅助蛋白功能

基本信息

批准号：
8316402
负责人：
SANDRA L QUACKENBUSH
金额：
$ 23.69万
依托单位：
COLORADO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-06-01 至 2014-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8316402
关键词：
Address Affect Animal Cancer Model Area Binding Biological Assay Boxing Cell Cycle Cell Cycle Progression Cell Cycle Regulation Cell Line Cell Proliferation Cell division Cell physiology Cells Complex Cyclin Gene Cyclins Dermal Development Disease Disease remission Dominant-Negative Mutation Elements Fishes Flow Cytometry Gene Expression Genes Genetic Transcription Goals Health Immune response In Vitro Interferons Investigation Knock-out Malignant Neoplasms Mediator of activation protein Modeling Molecular Molecular Cloning Mutate Mutation NF-kappa B Neoplasms Oncogenic Viruses Organism Outcome Phase Phospho-Specific Antibodies Phosphorylation Phosphotransferases Process Proteins Recombinants Research Retroviridae Role Simplexvirus Staging System Testing Therapeutic Intervention Transcript Transcription Coactivator Transcriptional Activation Transcriptional Regulation Tumor Suppressor Genes VP 16 Vertebrates Viral Viral Genes Viral Proteins Virus Virus Replication WDSV Work base cell growth cyclin C design functional outcomes gain of function mutation gene function human CDK3 protein human TAF9 protein loss of function mutation mutant neoplastic cell novel p65 promoter protein function research study sarcoma small hairpin RNA tissue/cell culture tumor tumor growth tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): Oncogenesis occurs when mechanisms of cell growth control are deregulated. This generally develops in a multistep process that involves mutations in oncogenes and tumor suppressor genes. The understanding of this process has been greatly enhanced through the study of tumor viruses. The piscine retroviruses and their associated neoplasias are new models of tumorigenesis. Walleye dermal sarcomas (WDS) are etiologically associated with the complex retrovirus, walleye dermal sarcoma virus (WDSV). These sarcomas develop and regress on a seasonal basis, providing a unique model to study molecular mechanisms of tumor development and regression in vertebrates. WDSV expression is highly regulated and the differential expression of viral transcripts correlates with the stage of disease development. Infectious virus is only produced during regression. Our working hypothesis is that the three WDSV accessory proteins, Orf A (rv-cyclin), Orf B, and Orf C, control viral gene expression and function in tumor growth and regression. WDSV rv-cyclin regulates transcription, inhibits virus expression, and enhances cell proliferation. Associations of rv-cyclin with cyclin dependent kinases 3 and 8 (cdk3 and cdk8), normally bound by host cyclin C, led to the hypothesis that rv-cyclin/cdk complexes alter viral and cellular gene expression and promote cell cycle progression. The long-term objective is to define novel mechanisms of tumor induction and control of gene expression. The proposed research has three specific aims: (1) Characterize rv-cyclin transregulation of transcription. The goals of this aim are to separate and define the mechanisms of rv-cyclin AD and cyclin box transcription control. These studies will focus on p53 and p53-responsive promoters that are regulated by both TAF9 and cyclin C/cdk8 and on NF-kB, which also contacts TAF9, and whose function is clearly inhibited by the rv-cyclin. (2) Define the functional outcome of rv-cyclin/cdk3 interaction. The identification of rv-cyclin-cdk3 interaction suggests a direct mechanism for cell cycle control by pRb phosphorylation. The kinase activity of rv-cyclin/cdk3 will be compared to cyclin C/cdk3, and the specific effects of the rv-cyclin on cell cycle regulation will be determined by flow cytometry. (3) Assess rv-cyclin function in control of virus expression and tumorigenesis. These studies will test wild type and mutated recombinant WDSV viruses with the rv-cyclin gene knocked out, with loss of function mutations in the cyclin box region and in the AD, and gain of function mutation of the AD. The replicative capacity of these infectious viruses will be tested in tissue culture cells and in fish, and the contribution of the rv-cyclin to tumor formation will be assessed. PUBLIC HEALTH RELEVANCE: The proposed research utilizes an efficient animal cancer model that combines tumor formation with natural and complete remission. The causative viral agent encodes a retroviral cyclin that controls cell growth and gene expression. These studies will identify targets for therapeutic intervention of cancer that are fundamental to all higher organisms.

描述（由申请人提供）：当细胞生长控制的机制放松调节时，就会发生肿瘤发生。这通常在多步过程中发展，该过程涉及癌基因和肿瘤抑制基因的突变。通过研究肿瘤病毒，对这一过程的理解大大增强了。 piscine逆转录病毒及其相关的肿瘤是肿瘤发生的新模型。角膜白斑皮肤肉瘤（WDS）在病因上与复杂的逆转录病毒，角膜白斑皮肤肉瘤病毒（WDSV）相关。这些肉瘤在季节性的基础上发展和退化，为研究脊椎动物中肿瘤发育和回归的分子机制提供了独特的模型。 WDSV表达受到高度调节，病毒转录本的差异表达与疾病发展阶段有关。传染病仅在回归过程中产生。我们的工作假设是，三种WDSV辅助蛋白ORF A（RV-CYCLIN），ORF B和ORF C，控制肿瘤生长和回归中的病毒基因表达和功能。 WDSV RV-Cyclin调节转录，抑制病毒表达并增强细胞增殖。 RV-Cyclin与细胞周期蛋白依赖性激酶3和8（CDK3和CDK8）的关联，通常由宿主细胞周期蛋白C结合，导致了RV-Cyclin/cdk复合物改变病毒和细胞基因表达并促进细胞周期进展的假设。长期目标是定义肿瘤诱导和控制基因表达的新机制。拟议的研究具有三个特定的目的：（1）表征转录的RV-CYCLIN转移。该目标的目标是分离和定义RV-Cyclin AD和Cyclin Box转录控制的机制。这些研究将集中于由TAF9和Cyclin C/CDK8和NF-KB调节的p53和p53响应启动子，它们也与TAF9接触，并且RV-Cyclin显然抑制了其功能。（2）定义RV-Cyclin/CDK3相互作用的功能结果。 RV-Cyclin-CDK3相互作用的鉴定表明通过PRB磷酸化来控制细胞周期的直接机制。 RV-Cyclin/CDK3的激酶活性将与细胞周期蛋白C/CDK3进行比较，RV-Cyclin对细胞周期调节的特定影响将通过流式细胞仪确定。（3）评估RV-CYCLIN在控制病毒表达和肿瘤发生中的功能。这些研究将用RV-Cyclin基因敲除野生型和突变的重组WDSV病毒，并且在Cyclin Box区域和AD中的功能突变损失，并获得AD的功能突变。这些感染性病毒的复制能力将在组织培养细胞和鱼类中进行测试，将评估RV-Cyclin对肿瘤形成的贡献。公共卫生相关性：拟议的研究采用了一种有效的动物癌模型，该模型将肿瘤形成与自然和完全缓解相结合。病毒药物编码控制细胞生长和基因表达的逆转录病毒细胞周期蛋白。这些研究将确定癌症治疗干预的靶标，这些靶标是所有更高生物基础的癌症。