RNA TARGETING WITH TERTIARY INTERACTIONS

描述：（改编自申请人的摘要）该R 03应用在 对题为创新药物发现研究的计划公告的回应 在艾滋病病毒感染方面。该项目的最终目标是 发现允许用小寡核苷酸靶向RNA的原理 基于抑制功能的化合物。制定这些原则可以 具有广泛的应用，包括设计治疗HIV的化合物， 机会性感染有待探索的新范式有：（1）约束性 三级相互作用增强（BETI），（2）自杀抑制，和（3） 剪接位点的多核苷酸催化水解。I组自我剪接 卡氏肺孢子虫核糖体RNA前体的内含子将作为 这些研究的模型系统。这种前体的正确拼接是 形成活性核糖体所必需的。初步研究表明， 核酸酶稳定，N3 '-P5'氨基磷酸酯六核苷酸显示结合 通过三级相互作用与序列紧密结合而增强， 通常与5 ′剪接位点对齐。这种六核苷酸抑制自我剪接 通过自杀抑制机制。拟议工作的具体目标是： (1)确定具有不带电荷的甲基膦酸酯键的寡核苷酸 表现出BETI和自我剪接的抑制。(2)确定BETI是否 在I组内含子的其他位点上可获得。(3)确定是否在体外 寡核苷酸抑制剂被卡氏肺孢子虫细胞摄取。结果 有望为设计许多RNA的抑制剂提供基本原则 功能协调发展的因为rRNA前体的成熟对于增殖是必不可少的 卡氏肺孢子虫，并且在人类基因中没有已知的I组自我剪接内含子， 该结果也可能为卡氏肺孢子虫肺炎提供潜在的治疗方法。

DESCRIPTION: (Adapted from Applicant's Abstract) This RO3 application is in response to a Program Announcement entitled Innovative Drug Discovery Research in AIDS Opportunistic Infections. The ultimate goal of this project is to discover principles that allow targeting of RNA with small oligonucleotide based compounds that inhibit function. Establishment of such principles could have wide ranging applications, including design of compounds to treat HIV and opportunistic infections. New paradigms to be explored are (1) binding enhancement by tertiary interactions (BETI), (2) suicide inhibition, and (3) oligonucleotide-catalyzed hydrolysis of splice sites. The group I self-splicing intron from the ribosomal RNA precursor in Pneumocystis carinii will serve as the model system for these studies. Correct splicing of this precursor is required for formation of active ribosomes. Preliminary work has shown that a nuclease stable, N3'-P5' phosphoramidate hexanucleotide exhibits binding enhanced by tertiary interactions in binding tightly to the sequence that normally aligns the 5' splice site. This hexanucleotide inhibits self-splicing via a suicide inhibition mechanism. Specific aims for the proposed work are: (1) Determine if oligonucleotides with uncharged methylphosphonate linkages exhibit BETI and inhibition of self-splicing. (2) Determine if BETI is available at other sites on the group I intron. (3) Determine if in vitro oligonucleotide inhibitors are taken up by P. carinii cells. The results are expected to provide fundamental principles for designing inhibitors of many RNA functions. Because maturation of rRNA precursor is essential for proliferation of P. carinii, and no group I self-splicing introns are known in human genes, the results could also provide potential therapeutics for P. carinii pneumonia.