ORGANIZATION AND EXPRESSION OF GENES IN VIRAL DNAS

腺病毒5型的EIA和EIB转录区域包含大约 3500个碱基对的DNA和编码功能，调节 其他病毒基因导致哺乳动物的致癌转化 细胞。该区域的DNA序列是已知的，其重叠的mRNAs 定义了它们的编码多肽并确定了其编码多肽。实验是 建议定义这些mRNA中的哪一个编码所需的多肽 无论是基因表达的调控还是细胞转化。 此外，这些编码多肽的功能结构域将是 确定了它们的作用机制，并对其作用机理进行了研究。规管 基因表达：I类宿主范围突变体定位基因产物 在EIA内表达其他病毒基因所需的； 方案将确定需要哪些特定的环评编码功能(S)。 真核转导载体将用于永久性猴的制备 含有野生型和突变型EIA和EIB区的细胞系 基因组。包含单个环评的全长拷贝的SV40病毒 将制备并使用cDNA来补充基因突变缺陷 永久细胞系。这将定义这些特定的基因功能 对于调节基因表达是必不可少的。它们的功能域 多肽将通过比较大量的EIA DNA序列来定义 I类寄主范围突变体和它们的回复突变体，并通过直接改变 通过体外诱变适当的cDNA的特定区域。这个 基因表达的调控机制将通过以下方式进行研究 高效生产环评多肽及利用富集物进行研究 它们与DNA的结合以及它们对基因体外转录的影响。 转换：将使用转换向量插入全长 将Ad5EIA和EIB中的DNA导入原代仓鼠细胞中，以确定 哪些基因产物可以诱导转化。寄主范围突变体和体外 突变将定义包含转化的结构域(S) 功能。此外，DNA高效的猿腺病毒7号也促进了 转化将被用来补充这些AD5研究。此数据 将作为实验研究其分子机制的基础。 哺乳动物细胞的致癌转化。

The EIA and EIB transcription regions of Adenovirus 5 contain approximately 3500 base pairs of DNA and encode functions that regulate the expression of other viral genes as result in the oncogenic transformation of mammalian cells. The DNA sequence of this region is known, its overlapping mRNAs defined and their encoded polypeptides determined. Experiments are proposed to define which of these mRNAs encode the polypeptides required for either the regulation of gene expression or cellular transformation. In addition the functional domains of these encoded polypeptides will be determined and their mechananism of action investigated. Regulation of Gene Expression: The class I host range mutants locate the gene products required for the expression of other viral genes within EIA; the following scheme will determine which specific EIA encoded function(s) are required. Eukaryotic transducing vectors will be used to prepare permanent monkey cell lines containing the EIA and EIB regions from wild type and mutant genomes. SV40 viruses containing full length copies of the individual EIA cDNAs will be prepared and used to complement the mutant defects in the permanent cell lines. This will define these specific gene functions essential for regulating gene expression. The functional domains of these polypeptides will be defined by comparing the EIA DNA sequences of numerous class I host range mutants and their revertants and by direct alteration of specific regions of the appropriate cDNAs by in vitro mutagenesis. The mechanisms by which gene expression is regulated will be studied by overproducing the EIA polypeptides and using enriched extracts to study their binding to DNA and their effect on in vitro transcription of genes. Transformation: Transducing vectors will be used to insert full length cDNAs from the Ad5 EIA and EIB into primary hamster cells to determine which gene products induce transformation. Host range mutants and in vitro mutagenesis will define the domain(s) containing the transformation functions. Also, simian Adenovirus 7 which is efficient in DNA promoted transformation will be used to complement these Ad5 studies. This data will serve as a basis for experiments to study the molecular mechanism of the oncogenic transformation of mammalian cells.