Diversity-Generating Retroelements in Phage and Bacterial Genomes

描述（由申请人提供）：我们发现了一个新的逆转录元件家族，称为多样性生成逆转录元件（DGR），其功能是使蛋白质编码DNA序列多样化。原型 DGR 在噬菌体基因组中被鉴定，其基础是它能够在基因中产生变异性，该基因指定博德特氏菌物种上受体分子的趋向性。博德特氏菌会引起人类和其他哺乳动物的呼吸道感染。向性转换是一种模板依赖性、逆转录酶介导的过程，它在目标基因内的指定位置引入核苷酸取代。这种基于盒的机制理论上能够在单个多肽中产生数万亿个不同的氨基酸序列，提供大量潜在的配体-受体相互作用。使用博德特氏菌噬菌体 DGR 作为特征，我们在许多细菌基因组中鉴定出了同源元件。特别值得注意的是，预计 DGR 可以使多形拟杆菌（胃肠道菌群中最丰富的成员之一）和齿垢密螺旋体（一种与牙周病相关的口腔细菌）表面的蛋白质多样化。原核 DGR 代表了一种产生蛋白质多样性的全新机制。除了作为新发现的遗传元件家族的基本重要性及其在人类健康和疾病中的潜在作用之外，DGR 还因其潜在应用而引起人们的兴趣。我们的具体目标是： 1. 对博德特氏菌噬菌体中存在的原型多样性生成逆转录因子进行机制分析。 2. 探索受体库和 DGR 编码受体蛋白配体识别的结构基础。 3. 研究人胃肠道共生多形拟杆菌中的 DGR 功能。

DESCRIPTION (provided by applicant): We have discovered a new family of retroelements, designated diversity-generating retroelements (DGRs) that function to diversify protein-encoding DNA sequences. The prototype DGR was identified in a bacteriophage genome on the basis of its ability to generate variability in a gene that specifies tropism for receptor molecules on Bordetella species. Bordetella cause respiratory infections in humans and other mammals. Tropism switching is a template-dependent, reverse transcriptase-mediated process that introduces nucleotide substitutions at defined locations within a target gene. This cassette-based mechanism is theoretically capable of generating trillions of different amino acid sequences in a single polypeptide, providing a vast repertoire of potential ligand-receptor interactions. Using the Bordetella phage DGR as a signature, we have identified homologous elements in numerous bacterial genomes. Of particular note are DGRs that are predicted to diversify proteins on the surface of Bacteroides thetaiotaomicron, one of the most abundant members of the gastrointestinal flora, and Treponema denticola, an oral bacterium associated with periodontal disease. Prokaryotic DGRs represent an entirely novel mechanism for generating protein diversity. In addition to their fundamental importance as a newly discovered family of genetic elements and their potential roles in human health and disease, DGRs are of interest as a result of their potential applications. Our specific aims are to: 1. Conduct a mechanistic analysis of the prototype diversity-generating retroelement present in Bordetella bacteriophage. 2. Probe the receptor repertoire and the structural basis of ligand recognition by a DGR-encoded receptor protein. 3. Investigate DGR function in the human gastrointestinal commensal Bacteroides thetaiotaomicron.