SEQUENCE AND STRUCTURE SIGNALS FOR 5S RNA PROCESSING IN DROSOPHILA MELANOGASTER

5S RNA，蛋白质所需的大核糖体亚基的一种小RNA 合成，具有保守的五个结构域的茎环二级结构。喜欢 大多数真核生物的RNA，D.Blackogaster 5S RNA不是直接在 抄写。135个核苷酸的初级转录本从其3‘端中去除了15个核苷酸 以处理结束。这项建议涉及5S RNA的结构基础 正在处理。 5S RNA的加工相对不受核苷酸变化的影响 加工点下游+120处，并从 阀杆IV-V另一方面，茎III中的缺失，移除了所有茎 IV-V，以及+1-118碱基对的破坏，第一对核苷酸 在茎I中，所有这些都干扰了加工。 在此，我建议彻底研究5S RNA所需的结构域 正在处理。与T7 RNA连锁的5S RNA基因的定点突变 将进行聚合酶启动子。突变的5S RNA将被转录 T7 RNA聚合酶，并用一只黑腹果蝇进行测试 加工萃取物用于其被加工的能力。 三个突变的集合将被引入到 螺旋茎打乱碱基配对，或进入顶部和底部以 恢复它。将进行成对的删除和插入，以更改 螺旋的长度。还将分析长度和顺序的影响 在每一个单股的环路中。通过这些实验，我们 应详细评估核苷酸序列的相对贡献 和碱基配对结构对果蝇加工能力的影响 黑素胃泌素5S RNA。

5S RNA, a small RNA of the large ribosomal subunit required for protein synthesis, has a conserved five domain stem-loop secondary structure. Like most eukaryotic RNAs, D. melanogaster 5S RNA is not utilized directly after transcription. The 135 nt primary transcript has 15 nt removed from its 3' end by processing. This proposal concerns the structural basis for 5S RNA processing. 5S RNA processing is relatively unaffected by changes in the nucleotides downstream from the processing site at +120, and by deletion of 30 nt from stem IV-V. On the other hand, deletions in stem III, removing all of stem IV-V, and disruption of the +1-118 base pair, the first paired nucleotides in stem I, all interfere with processing. Herein I propose to thoroughly investigate the domains required for 5S RNA processing. Site-directed mutagenesis of 5S RNA genes linked to a T7 RNA polymerase promoter will be performed. Mutant 5S RNAs will be transcribed with T7 RNA polymerase and tested using a Drosophila melanogaster processing extract for their ability to be processed. Sets of three mutations will be introduced into the top or bottom of helical stems to disrupt base pairing, or into both top and bottom to restore it. Paired deletions and insertions will be made to alter the length of helixes. The effect of length and sequence will also be analyzed in each of the single stranded loops. By means of these experiments, we shall evaluate in detail the relative contribution of nucleotide sequence and base paired structure to the processing ability of Drosophila melanogaster 5S RNA.