FACTORS INVOLVED IN TRANSCRIPTION BY RNA POLYMERASE II

本项目的目的是为了更好地了解 RNA聚合酶II转录的延伸期。 形成 和维持能够合成 将研究mRNA。 这项工作的主要目标之一是 描述了两个最近发现的似乎与 控制这种复合物的形成。 其中一个因素是 与前起始复合物紧密偶联，并迫使聚合酶进入 一种仅能形成短转录物的延伸模式。 的 第二个因子在起始后起作用，并允许聚合酶继续进行， 转录基因的3'端。 一种体外方法， 果蝇Kc细胞核提取物将被用于所有的建议。 问题研究 固定在顺磁珠上的DNA模板将用于 操纵在生理启动子上形成的转录复合物。 导致成熟伸长形成的过程的解剖 复合物将通过改变存在于 在络合物的磁性浓缩之后的反应。 一个 对成熟伸长复合体的性质的检查将确定 延伸因子与复合物相互作用的稳定性， 由不同启动子引发形成的延伸复合物具有 不同的属性。 另一个目标是确定 DmS-II是一种果蝇RNA聚合酶II， 因子 这些研究将有助于获得大量的 完整的蛋白质和蛋白质的结构域已经产生， E. coli中获得。 延伸率控制的明显参与， 许多癌基因和病毒（包括HIV）的表达，要求 该过程的生物化学细节被阐明，以便合理地 可以采取控制疾病状态的方法。 拟议 研究应该阐明控制伸长的机制， 此外，将允许形成一个更连贯的画面， 完整的转录周期。

The goal of this proposed project is to come to a better understanding of the elongation phase of transcription by RNA polymerase II. The formation and maintenance of elongation complexes which are competent to synthesize mRNAs will be investigated. One of the major goals of the work is to characterize two recently identified factors that seem to be involved in controlling the formation of such complexes. One of these factors is tightly coupled to the preinitiation complex and forces the polymerase into an elongation mode in which only short transcripts can be formed. The second factor acts after initiation and allows the polymerase to proceed to the 3' end of the transcribed gene. An in vitro approach utilizing Drosophila Kc cell nuclear extracts will be taken for all of the proposed studies. DNA templates immobilized to paramagnetic beads will be used to manipulate transcription complexes formed on physiological promoters. Dissection of the processes leading to the formation of mature elongation complexes will be accomplished by changing the factors present in the reactions following magnetic concentration of the complexes. An examination of the properties of mature elongation complexes will determine how stably the elongation factors interact with the complex and if elongation complexes formed by initiation from different promoters have different properties. Another goal is to determine the biochemical details of the function of DmS-II, a Drosophila RNA polymerase II elongation factor. These studies will be aided by the availability of large amount of the intact protein and domains of the protein which have been produced in E. coli from the cDNA. The clear involvement of elongation control in the expression of numerous oncogenes and viruses including HIV, demands that the biochemical details of the process be elucidated so that rational approaches to controlling the diseased state may be made. The proposed studies should shed light on the mechanisms controlling elongation and, furthermore, will allow the formation of a more coherent picture of the complete transcription cycle.