DEVELOPING AN IN VIVO MODEL OF THE HUMAN PAPILLOMAVIRUS E5 ONCOGENE

该子项目是利用资源的许多研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其他NIH来源。 为子项目列出的总成本可能 代表子项目使用的中心基础设施的估计数量， NCRR赠款不直接向子项目或子项目工作人员提供资金。 侵袭性宫颈癌和前体病变与人乳头瘤病毒（HPV）感染（尤其是PHV 16型）的存在密切相关。 HPV 16表达三种癌基因- E5、E6和E7。 由E6和E7癌基因编码的蛋白质已显示分别抑制肿瘤抑制基因p53和Rb的功能（CITE）。 尽管有大量证据表明E5的表达可以引起细胞转化，但其发生的机制一直存在争议。 我们最近发现HPV 16 E5蛋白的表达可以诱导细胞融合，导致双核细胞，核增大和四倍体。 有趣的是，这些都是癌性和前驱病变的特征。 基于这些发现和其他人的工作，我们假设E5通过诱导细胞-细胞融合和随后的染色体含量不稳定来启动肿瘤发生。 然而，这些融合事件背后的分子机制仍然不清楚。 我们建议验证和扩展我们的体外研究结果，通过开发和体内模型的HPV E5功能，果蝇作为一个模式生物。 我们将开发一个转基因果蝇模型来表征HPV 16 E5表达的表型后果。 我们将首先研究果蝇发育过程中普遍存在的HPV 16 E5表达的影响。 接下来，我们将在果蝇上皮细胞中特异性表达HPV 16 E5，以测试PHV 16 E5表达是否能诱导与癌症相关的异常细胞形态。 这些实验将有助于建立HPV 16 E5基因的体内模型，这将使未来的工作朝着阐明HPV诱导的致癌作用的遗传和细胞决定因素。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Invasive cervical carcinoma and precursor lesions are strongly associated with the presence of human papillomavirus (HPV) infection, particularly the PHV16 type. HPV16 expresses three oncogenes - E5, E6, and E7. The proteins encoded by the E6 and E7 oncogenes have been shown to inhibit the function of the tumor suppressor genes p53 and Rb, respectively (CITE). Despite numberous lines of evidence that expression of E5 can cause cell transformation, the mechanism by which this occurs has been controversial. We have recently discovered that the expression of HPV16 E5 protein can induce cell-cell fusion resulting in bi-nucleated cells, enlarged nuclei, and tetraploidy. Interestingly, these are all characteristics of cancerous and precursor lesions. Based on these findings and the work of others, we hypothesize that E5 initiates tumorigenesis by inducing cell-cell fusion and subsequent instability in chromosome content. HOwever, the molecular mechanisms that underlie these fusion events are still unclear. We propose to verify and extend our in vitro findings by developing and in vivo model of HPV E5 function using Drosophila melanogaster as a model organism. We will develop a transgenic Drosophila model to characterize the phenotypic consequences of HPV16 E5 expression. We will first look at the effect of ubiquitous HPV16 E5 expression during Drosophila development. Next, we will express HPV16 E5 specifically in Drosophila epithelial cells i vivo to test whether PHV16 E5 expression can induce aberrant cell morphologies thatare associated with cancer. These experiments will help to establish an invivo model of the HPV16 E5 gene that will allow future work toward elucidating the genetic and cellular determinants of HPV-induced carcinogenesis.