MECHANISTIC APPROACHES TO HSV-2 INDUCED TRANSFORMATION

单纯疱疹病毒2型(HSV-2)是一种大型DNA病毒， 感染人类，主要通过性接触传播。 宫颈癌最常发生在乱交的妇女中 或者是在有乱交伴侣的女性身上。此外，HSV-2 在女性中检测到抗原和核酸的频率更高 患有宫颈癌的人比没有患宫颈癌的人更容易患上宫颈癌。因此，HSV-2可能是 要么是宫颈癌的病因，要么是几种 表现转化状态所需的辅因子。这个 单纯疱疹病毒2型BamHI-E片段的致癌作用 已建立的啮齿动物细胞的转化和最小 BamHI-E的转换区(MtrIII)映射到486个碱基对 碎片。DNA序列基序介导的转化 在MTRIII中，它类似于控制重组的元件和 转录，而不是病毒蛋白的表达。这个 本提案中提出的问题旨在勾勒出 HSV-2诱导转化的机制。做细胞原形- 癌基因在HSV-2介导的转化中起作用？至 回答这个问题，核糖核酸的稳态水平 转化细胞将与正常细胞中的细胞进行比较。做 转录调控序列在mtrIII中起着关键作用。 在HSV-2介导的转化和基因表达中？一个 细菌氯霉素乙酰转移酶报告基因 基因将被用来关联转录增强子 MtrIII的活性与转化及HSV-2的表达 基因。DO潜在的茎环结构和DNA构象 在MTRIII中介导的肿瘤转化？特定于站点 MTRIII中茎环结构的诱变及其作用 我们将分析这类突变对转化的影响。各地区 已经改变DNA构象的MTRIII将被绘制成图谱 使用特定的化学探针，并与 转型。HSV-2转化的细胞是否保留mtrIII DNA 序列呢？将对转换的细胞进行分析，以整合 单纯疱疹病毒2型的Southern blotting实验和上体测序 含有HSV-2 DNA的成分将被识别和 在转化的细胞中进行分析。单纯疱疹病毒2型具有转化功能吗？ 它的自然宿主--人类的潜力？转型 将在几个人类细胞系中检测HSV-2的潜力。在……里面 证据表明单纯疱疹病毒2型是一种 宫颈癌，这些研究将定义一种结构/功能 一种新的转化域与细胞鉴定的关系 在转化过程中发生改变的基因。

Herpes simplex virus type 2 (HSV-2) is a large DNA virus which infects humans and is spread primarily through sexual contact. Cervical carcinoma occurs most frequently in promiscuous women or in women with promiscuous partners. Furthermore, HSV-2 antigens and nucleic acids are detected more frequently in women with cervical cancer than in those without. Thus, HSV-2 may be either the causative agent of cervical cancer or one of several cofactors required to manifest the transformed state. The BamHI-E fragment of HSV-2 induces the neoplastic transformation of established rodent cells and the minimal transforming region of BamHI-E (mtrIII) maps to a 486 base pair fragment. Transformation is mediated by DNA sequence motifs in mtrIII which resemble elements controlling recombination and transcription and not the expression of a viral protein. The questions asked in this proposal are intended to delineate the mechanism of HSV-2 induced transformation. Do cellular proto- oncogenes play a role in HSV-2 mediated transformation? To answer this question, the steady state levels of RNA in transformed cells will be compared to that in normal cells. Do the transcriptional regulatory sequences in mtrIII play a key role in HSV-2 mediated transformation and gene expression? A reporter gene, the bacterial chloramphenicol acetyltransferase gene, will be utilized to correlate the transcriptional enhancer activity of mtrIII with transformation and the expression of HSV-2 genes. Do potential stem-loop structures and DNA conformation in mtrIII mediate neoplastic transformation? Site-specific mutagenesis of a stem-loop structure in mtrIII and the effects of such mutations on transformation will be analyzed. The regions of mtrIII that have altered DNA conformation will be mapped using specific chemical probes and correlated with transformation. Do cells transformed by HSV-2 retain mtrIII DNA sequences? Transformed cells will be analyzed for integrated HSV-2 sequences by Southern blotting experiments and episomal elements which contain HSV-2 DNA will be identified and analyzed in transformed cells. Does HSV-2 posses transforming potential in its natural host, the human? The transforming potential of HSV-2 will be assayed in several human cell lines. In light of evidence which implicates HSV-2 as a causative agent of cervical cancer, these studies will define a structure/function relationship of a novel transforming domain and identify cellular genes that are altered during transformation.