HIV-2 GENOMIC RNA STRUCTURE

考虑到逆转录病毒编码的复制控制的复杂性，逆转录病毒的基因组非常小。为了达到这种程度的简约性，基因组RNA不仅包含蛋白质的编码信息，而且还包含反式激活、二聚化、双核苷化、剪接和靠近5'和3'末端的多聚腺苷酸添加的功能信号。这些事件中的每一个都对病毒的有效复制至关重要。在这里提出的研究中，将在HIV- 2病毒（人类艾滋病的病原体）的背景下详细研究基因组RNA中促进全长基因组两个拷贝二聚化的顺式元件的结构和功能。具体而言，将使用RNA结构探测、化学修饰干扰分析、诱变和缺失分析来鉴定和表征基因组RNA二聚化的信号。通过这些实验确定的位点将使用生物化学方法和组合体外进化方法进一步表征，以确定相互作用的稳定性和机制。将构建基因组RNA的5'前导区和部分gag编码序列的详细二级/三级结构图，以更好地理解该区域中功能基序之间的潜在相互作用。最后，将构建在其二聚化结构域中具有缺失或突变的突变体病毒，以评估其体内作用。这些研究将为开发针对基因组RNA功能区域的新抗逆转录病毒疗法提供重要的结构基础。

The genomes of retroviruses are remarkably small considering the complexity of replicative control which they encode. To attain this level of parsimony, the genomic RNA contains not only coding information for proteins, but also functional signals for transactivation, dimerization, encapsidation, splicing, and poly A addition near the 5' and 3' extremities. Each of these events is vital for productive replication of the virus. In the research proposed here, the structure and function of the cis-element(s) within the genomic RNA that promote dimerization of two copies of the full-length genome will be studied in detail within the context of the HIV- 2 virus, a causative agent for AIDS in humans. Specifically, the signal for dimerization of genomic RNA will be identified and characterized using RNA structure probing, chemical modification interference analysis, mutagenesis, and deletion analysis. Sites identified by these experiments will be further characterized using biochemical methods and a combinatorial in vitro evolution approach to determine the stability and mechanism of the interaction. Detailed secondary/tertiary structure maps of the 5' leader region and parts of the gag coding sequence of the genomic RNA will be constructed to better understand potential interplay between functional motifs in the region. Finally, mutant viruses will be constructed with deletion or mutations in their dimerization domains to assess their effects in vivo. These studies will provide a vital structural basis for developing new anti-retroviral therapies directed toward the functional regions of the genomic RNA.