YEAST MRNA 3' END FORMATION AS AN ANTIFUNGAL DRUG TARGET

多聚腺苷酸化3'末端的形成是多聚腺苷酸化3'末端的形成中的重要步骤。 真核生物mRNA的生物合成。在哺乳动物中， 广泛地在体内，以及在无细胞系统中，并已确定 多聚腺苷酸化的特定序列。在低等真核生物中， 酵母，不到50%的mRNA检查到目前为止，有一个可识别的 多聚腺苷酸化信号等同于哺乳动物序列。我们相信， 因此，真菌中mRNA 3'末端的形成可能是通过 与哺乳动物中发生的机制显著不同 细胞这个处理步骤可以代表一个新的目标， 抗真菌药物的开发。在第一阶段，我们将构建3个报告器 质粒，以确认酵母多聚腺苷酸化信号不同于 哺乳动物。缺失突变和体外聚腺苷酸化系统 将用于定义酵母多聚腺苷酸化信号序列。作为 第二阶段的一部分，我们将开发一个体内系统与记者 质粒来筛选抑制酵母中3'末端形成的化学品。 同样在第二阶段，体外系统将用于克隆酶 参与3'末端形成，并证实抑制3'末端形成的化学物质 体内系统实际上抑制3'末端形成。

The formation of polyadenylated 3' termini is an essential step in the biogenesis of eukaryotic mRNA. In mammals, this process has been examined extensively in vivo, as well as in cell-free systems, and has identified specific sequences for polyadenylation. in lower eukaryotes such as yeast, less than 50% of the mRNAs examined to-date have an identifiable polyadenylation signal equivalent to the mammalian sequence. We believe, therefore that the formation of mRNA 3' ends in fungi may proceed by a mechanism significantly different than that which occurs in mammalian cells. This processing step could represent a novel target for the development of antifungal drugs. In Phase I we will construct 3 reporter plasmids to confirm that yeast polyadenylation signals differ from that of mammals. Deletion mutagenesis and an in vitro polyadenylation system will be used to define the yeast polyadenylation signal sequences. As part of the Phase II we will develop an in vivo system with the reporter plasmids to screen chemicals for inhibition of 3' end formation in yeast. Also in Phase II the in vitro system will be used to clone enzymes involved in 3' end formation and confirm that chemicals inhibiting in vivo systems actually inhibit 3' end formation.