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Spatiotemporal Gene Regulation and KSHV Replication

时空基因调控和 KSHV 复制

基本信息

批准号：
10436841
负责人：
Yoshihiro Izumiya
金额：
$ 35.91万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-07-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10436841
关键词：
3-Dimensional Address Architecture Awareness B-Cell Lymphomas Bacterial Artificial Chromosomes Binding Binding Sites Biochemical Biochemical Genetics Biological Process Cell Nucleus Cells Chromatin Structure Chromosomes Code Complex Development Disease Electron Microscopy Electrons Elements Enzymes Episome Escherichia coli Fluorescence Microscopy Gene Activation Gene Expression Gene Expression Regulation Genes Genetic Genetic Transcription Genome Genomic Segment Genomics Goals Hi-C Human Herpesvirus 8 Image Immunoprecipitation In Situ Incidence Infection Informatics Investigation Kaposi Sarcoma Knowledge Lead Light Link Malignant Neoplasms Mass Spectrum Analysis Mediating Methods Microscopy Modeling Molecular Molecular Genetics Multicentric Angiofollicular Lymphoid Hyperplasia Mutation Nuclear Optics Poly A Process Promoter Regions Property Proteins RNA RNA Binding RNA Polymerase II Recombinants Regulator Genes Research Resolution Role Series Singlet Oxygen Small Interfering RNA Structure System Therapeutic Time Transcript Transcriptional Regulation United States National Institutes of Health Untranslated RNA Viral Viral Genes Viral Genome Viral Proteins Virus Assembly Virus Replication base cellular imaging chromosome conformation capture deep sequencing genetic analysis genetic approach genomic RNA insight latency-associated nuclear antigen live cell microscopy microscopic imaging mutant novel therapeutic intervention physical property primary effusion lymphoma programs promoter protein K recruit spatiotemporal tool transcription factor viral genomics virology

项目摘要

Kaposi's sarcoma-associated herpesvirus (KSHV) genomic episomes dynamically move in three- dimensional (3D) nuclear space to ultimately aggregate in reactivating cells. Cellular RNA polymerase II (RNA pol II) also translocates onto the aggregated viral genomes, resulting in the formation of viral transcriptional factories. This previously uncharacterized mechanism represents an additional layer of KSHV gene regulation, which facilitates the reutilization and repurposing of the limited quantities of cellular RNA pol II for effective viral gene expression. Understanding this mechanism may allow us to identify new targets to inhibit viral replication. In this application, we propose a comprehensive study to (1) identify the key cellular proteins involved with viral transcriptional factory structure or function; (2) determine the role of noncoding PAN RNA transcripts and its genomic region in viral transcriptional factory formation; and (3) visualize the physical assembly of viral transcriptional factories in the nuclei of reactivated host cells using live 3D fluorescence microscopy. We will also characterize the molecular architecture of viral transcriptional factories by super-resolution optical microscopy with correlative electron microscopic imaging. Through this multifaceted approach, we hope to understand the role of viral transcriptional factories in the amplification of KSHV gene expression, which may in turn lead to the development of new therapeutic strategies for KSHV-associated malignancies by inhibiting viral replication. In addition, we expect this investigation will shed light on the spatio-temporal (4D) organization of the cell nucleome: using KSHV as a well-defined model chromosome that we can readily track in the host cell nucleus with biochemical and genetic approaches.

卡波西肉瘤相关疱疹病毒(KSHV)基因组上游体在3个月内动态移动三维(3D)核空间最终聚集在重新激活的细胞中。细胞RNA聚合酶II(RNA Pol II)也转位到聚集的病毒基因组上，导致形成病毒转录工厂。这一以前未被描述的机制代表了KSHV基因调控的额外一层，它促进了有限数量的细胞RNA polII的再利用和再利用，以获得有效的病毒基因表达。了解这一机制可能会让我们找到新的靶点来抑制病毒复制。在这一应用中，我们建议进行一项全面的研究，以(1)确定涉及的关键细胞蛋白具有病毒转录工厂结构或功能；(2)确定非编码PAN RNA转录本的作用及其基因组区域在病毒转录工厂的形成；以及(3)可视化病毒的物理组装用实时3D荧光显微镜观察重新激活的宿主细胞的细胞核中的转录工厂。我们会也用超分辨光学方法表征病毒转录工厂的分子结构显微镜结合相关电子显微镜成像。通过这种多方面的方法，我们希望了解病毒转录工厂在KSHV基因表达扩增中的作用，这可能在转而通过抑制病毒来开发治疗KSHV相关恶性肿瘤的新策略复制。此外，我们预计这项研究将有助于揭示时空(4D)组织细胞核组：使用KSHV作为明确的模式染色体，我们可以很容易地在宿主细胞中追踪到它用生物化学和遗传学的方法来研究细胞核。