Human Placental CMV Infection: Global Gene Expression

描述（由申请人提供）：本申请的研究旨在利用DNA微阵列技术全面了解胎盘中分化细胞滋养细胞（CTBs）的基因表达，并评估人巨细胞病毒（CMV）感染对这些细胞表达途径的影响。我们试图通过鉴定胎盘发育过程中CTBs分化失调的病毒基因来定义CMV发病机制的新生物学。通过与DNA微阵列技术先驱Joseph DeRisi博士的实验室合作，我们建议大幅扩大亲本资助（AI46657）的范围。我们的计划是使用人类和巨细胞病毒基因组阵列来探索巨细胞病毒感染前后CTBs中的基因表达途径。这种方法将允许对整个转录程序进行询问，而不需要先验地选择要研究的基因。因此，发现新的或意想不到的基因表达的重要变化的可能性非常高，如果没有这项新技术实现的全球评估，这将是非常困难的。DeRisi博士在UCSF为基因组学和蛋白质组学组织了一个微阵列核心设施，提供微阵列打印、杂交和数据分析方面的设备、协议、软件和技术培训。在DeRisi博士实验室的帮助下，我们将在包含70个聚核苷酸的微阵列上同时检测细胞和病毒基因表达模式，这些寡核苷酸代表了14000个人类基因和整个巨细胞病毒基因组。我们的具体目标是：(1)确定细胞基因表达模式在CTBs分化中的作用，并研究CMV感染对实验室和致病菌株的影响；(2)鉴定影响CTB表达途径的CMV基因。微阵列实验中提示的表达变化将在父母资助的支持下，通过蛋白质和RNA水平分析和功能分析独立验证。通过此次合作获得的专业知识和资源将使我们能够全面了解与CTB分化和免疫功能相关的表达途径，并加深我们对CMV在临床相关原代细胞类型中的发病机制的理解。微阵列技术促进的基因探索可以揭示分子诊断的新靶点，并导致阻止产前巨细胞病毒感染的治疗。CTBs和致瘤细胞中与侵袭和血管生成相关的表达途径之间的相似性表明，这些研究在确定治疗新靶点和CMV基因方面具有额外的潜力，这些靶点可用于治疗癌症中的肿瘤进展。

DESCRIPTION (provided by applicant): Studies in this application are designed to use DNA microarray technology to attain a comprehensive understanding of gene expression in differentiating cytotrophoblasts (CTBs) in the placenta and to assess the impact of human cytomegalovirus (CMV) infection on expression pathways in these cells. We seek to define a new biology of CMV pathogenesis by identifying viral genes that dysregulate the differentiation of CTBs during placental development. Through a collaboration with the laboratory of Dr. Joseph DeRisi, a pioneer in DNA microarray technology, we propose to broaden the scope of the parent grant (AI46657) substantially. Our plan is to use human and CMV genomic arrays to explore gene expression pathways in CTBs before and after CMV infection. This approach will allow interrogation of the entire transcriptional program without requiring an a priori choice of which genes to study. Thus, there is a very high likelihood of discovering important changes in the expression of novel or unexpected genes, which would be very difficult without the global assessment made possible by this new technology. Dr. DeRisi has organized a microarray core facility for genomics and proteomics at UCSF that offers equipment, protocols, software, and technical training in microarray printing, hybridization, and data analysis. With assistance from the Dr. DeRisi laboratory, we will examine patterns of cellular and viral gene expression simultaneously on microarrays containing 70-mer oligonucleotides representing 14,000 human genes and the entire CMV genome. Our specific aims are (1) to determine patterns of cellular gene expression in differentiating CTBs and investigate the impact of CMV infection with laboratory and pathogenic strains and (2) to identify CMV genes that affect CTB expression pathways. Changes in expression suggested in microarray experiments will be verified independently by analysis of protein and RNA levels and functional assays with support of the parent grant. The expertise and resources available through this collaboration will enable us to achieve a global view of expression pathways associated with CTB differentiation and immune functions and to deepen our understanding of CMV pathogenesis in a clinically relevant primary cell type. Gene exploration facilitated by microarray technology could reveal novel targets for molecular diagnosis and lead to therapies that block prenatal CMV infection. Similarities between expression pathways associated with invasion and angiogenesis in CTBs and tumorigenic cells suggest the added potential of these studies to identify novel targets for therapies and CMV genes that could be used to treat tumor progression in cancer.