MOLECULAR BIOLOGY OF THE DELTA AGENTS

我们建议再进行五年的研究， 人丁型肝炎病原体的性质及其对人丁型肝炎的影响 环状基因组RNA分子，使其在病毒和 细胞RNA。 δ RNA仍然是唯一的非编码基因组RNA， 利用细胞组分，通过 滚环机制迄今已在哺乳动物细胞中发现。 的 RNA水平的事件，这种代理带来的承担其复制 和宿主细胞的相互作用包括RNA催化的酶促事件 在复制过程中使用内置核酶切割RNA; 后代RNA的特异性RNA连接以形成环;特异性RNA 利用宿主酶编辑mRNA阅读框;广泛 二级和局部三级结构，包括特征紫外线- 在基因组和互补的可交联结构 在复制过程中产生的反基因组链;相互作用 与宿主蛋白激酶PKR在特定的RNA位点结合; 宿主RNA聚合酶分子的位点（RNA启动子）。 许多 这些过程和相互作用涉及保守的、非编码的 delta RNA区域，其中一些部分涉及多个 需要构象转换的特定相互作用。 研究δ基因组相互作用是我们的长期目标 和反基因组RNA与宿主细胞成分;并了解如何 这些相互作用和基于RNA的活动联合收割机 高效地复制这种小型的、圆形的、高度结构化的 导致严重人类疾病的单链RNA因子。 我们 具体目的是（i）通过携带病毒来研究δ病毒的发病机制， 研究特定的宿主蛋白delta RNA相互作用， 强调蛋白激酶和RNA编辑活动;（ii）由于 delta的RNA：蛋白质相互作用使用特定的蛋白质和RNA 具有普遍生物学意义的图案，我们的目标是追求 结合蛋白质的delta RNA关键元件的详细结构 并以其他方式反应以促进复制和发病， 物理和生物化学技术;以及（iii）通过 RNA酶保护等手段，将RNA水平的酶 负责复制链的切割和连接， 它们通过RNA水平上的构象转换进行调节。

We propose to conduct an additional five years of research on the properties of the causative agent for human delta hepatitis and its circular genomic RNA molecular which make it unique among viral and cellular RNAs. Delta RNA remains the only non-coding genomic RNA to undergo efficient RNA-to-RNA copying using cellular components by a rolling circle mechanism so far identified in mammalian cells. The RNA-level events which this agent brings to bear on its replication and host cell interactions include RNA-catalyzed enzymatic events using built in ribozymes for RNA cleavage during replication; specific RNA ligation of progeny RNAs to form circles; specific RNA editing of a mRNA reading frame utilizing a host enzyme; extensive secondary and local tertiary structure, including characteristic UV- crosslinkable structures in both the genomic and complementary antigenomic strands which arise during replication; interaction(s) with a host protein kinase, PKR, at specific RNA sites; and binding sites (RNA promoters) for host RNA polymerase molecules. Many of these processes and interactions involve the conserved, non-coding region of delta RNA, some parts of which are involved in multiple specific interactions requiring conformational switching. It is our long-term goal to study the interactions of delta genomic and antigenomic RNAs with host-cell components; and to understand how these interactions and RNA-based activities combine to promote efficient replication of this small, circular highly structured single-stranded RNA agent which causes a serious human disease. Our specific aims are (i) to study delta viral pathogenesis by carrying out studies on specific host protein delta RNA interactions, with emphasis on the protein kinase and RNA editing activities; (ii) since delta's RNA:protein interactions employ specific protein and RNA motifs of great general biological interest, we aim to pursue the detailed structure of key elements in delta RNA which bind proteins and otherwise react to promote replication and pathogenesis, using physical as well as biochemical techniques; and (iii) to study, by RNase protection and other means, the RNA-level enzymes of delta responsible for cleavage and ligation of replicating strands and their regulation by conformational switching at the RNA level.