Genetic Mechanisms Driving Evolution of Rickettsiaceae

描述（由申请人提供）：拟议研究的总体目标是显著提高我们目前对立克次体性质的理解，立克次体是一组专性细胞内节肢动物传播的细菌，在世界不同地区引起至少11种人类疾病。恙虫病东方体（Orientia tsugamushi）、赤立立克次体（Rickettsia akari）、R. canada和R.贝利将被研究。这些生物体的完整基因组序列将被确定为99.99%的准确度，并通过计算机进行分析，以识别每种生物体中存在的推定RNA、蛋白质编码基因、假基因、重复元件、“自私DNA”和调控元件。这些信息将在可公开查阅的数据库中提供。在每一个立克次体中发现的独特和保守的遗传元件将进行比较，以帮助了解其致病性适应进化的机制。这种最初的计算机分析需要生物化学确认，因为它取决于立克次体基因序列与其他细菌中发现的具有良好特征的遗传元件的相似性。在这些物种中预期存在的1000-2000个基因中，只有不到15个基因以前被研究过。由于立克次体的细胞内生活方式和缺乏遗传系统，传统的方法来表征这些基因在立克次体的功能特性是有限的。将为每个立克次体基因组构建包含每个基因的细菌人工染色体（BAC）文库和DNA阵列。BAC和DNA阵列将用于表征立克次体基因的表达和调控。将克隆和表达可能用于疫苗开发和作为潜在治疗靶点的选定立克次体蛋白，以促进其表征。

DESCRIPTION (provided by applicant): The overall goal of the proposed research is to significantly enhance our present understanding of the properties of rickettsiae, a group of obligately intracellular, arthropod transmitted, bacteria which cause at least eleven human diseases in different parts of the world. Orientia tsutsugamushi, Rickettsia akari, R. canada, and R. bellii will be studied. The complete genome sequences of these organisms will be determined to 99.99% accuracy and analyzed by computer to identify the putative RNA, protein encoding genes, pseudogenes, repeat elements, 'selfish DNA', and regulatory elements present in each organism. This information will be provided in publicly accessible databases. Unique and conserved genetic elements found in each rickettsia will be compared to help understand the mechanisms underlying evolution of their pathogenic adaptations. This initial in silico analysis requires biochemical confirmation as it depends upon the similarity of rickettsial gene sequences to well-characterized genetic elements found in other bacteria. Less than 15 of the 1000-2000 genes expected to be present in each of these species have been studied previously. Because of the intracellular lifestyle of rickettsiae and the lack of genetic systems, conventional approaches to characterization of the functional properties of these genes in rickettsiae are limited. Bacterial artificial chromosome (BAC) libraries and DNA arrays containing each gene will be constructed for each rickettsial genome. The BACs and DNA arrays will be used to characterize the expression and regulation of rickettsial genes. Selected rickettsial proteins that may be useful in vaccine development and as potential therapeutic targets will be cloned and expressed to facilitate their characterization.