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Role of selfish restriction-modification gene complexes in genome evolution : comparative genomics and experimental evolution

自私限制修饰基因复合物在基因组进化中的作用：比较基因组学和实验进化

基本信息

批准号：
15370099
负责人：
KOBAYASHI Ichizo
金额：
$ 9.79万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

1.Conditions for post-segregational host killing (genetic addiction)We found that EcoRI modification enzyme is as stable as EcoRI restriction enzyme and bulk cellular proteins. We analyzed conditions for evolution of addiction genes using evolutionary ecology approach. Importance of space structure was demonstrated.2.Restriction enzyme and modification enzyme obtained through comparative genomics with hyperthermophilic archaeaThrough comparison of the genomes of Pyrococcus abyssi and Pyrococcus horikoshii, we identified a putative restriction-modification gene complex inserted into the former genome. We identified restriction enzyme PabI through expression screening in wheat germ-derived cell-free protein synthesis system. It generates a novel terminus TA3' at 5'GTAC.3.Reconstruction of formation of large genome polymorphisms through comparison of closely-related genomesComparing 7 genomes of Staphylococcus aureus, we found evolution of S (specificity) subunit of Type I restriction enz … More yme through combination of two target recognition domains. From analysis of three tandem paralogue gene clusters, we concluded that some genome rearrangments have resulted from homologous recombination between sequence within each unit conserved for its functional importance. Comparing 4 genomes of Neisseria, we identified filamentous phages likely integrated by its transposase and occasionally rearranged by it.4.DNA breakage and genome rearrangements triggered by restriction breaks and othersWe found that Type III restriction breakage after single infection of bacteriophage lambda is repaired by homologous recombination function of prophage. Homology-driven illegitimate recombination was found to depend on RecA and Type I restriction sites. We developed a sensitive assay of linearization of circular bacterial chromosome and analyzed various recombination-defective mutants of E.coli. We demonstrated high level of DNA double-strand break repair in BJ5183 strain of E.coli. We modeled asymmetric branch migration of Holliday structure and analyzed data in vitro.5.Multiplication of a restriction-modification gene complexWe showed that tandem amplification of BamHI restriction-modification gene complex on Bacillus subtilis chromosome is dependent on function of the restriction enzyme. Less

1.分离后杀死宿主的条件（遗传成瘾）我们发现EcoRI修饰酶与EcoRI限制性酶和大量细胞蛋白一样稳定。我们使用进化生态学方法分析了成瘾基因的进化条件。证明了空间结构的重要性。2.通过与超嗜热古菌的比较基因组学获得的限制性酶和修饰酶通过比较深渊火球菌和堀越火球菌的基因组，我们鉴定了一个插入到前基因组中的假定的限制性修饰基因复合物。我们通过在小麦胚芽衍生的无细胞蛋白质合成系统中的表达筛选鉴定了限制性内切酶PabI。它在 5'GTAC 处生成了一个新的末端 TA3'。 3.通过比较密切相关的基因组来重建大基因组多态性的形成通过比较金黄色葡萄球菌的 7 个基因组，我们通过两个目标识别域的组合发现了 I 型限制性酶的 S（特异性）亚基的进化。通过对三个串联旁系同源基因簇的分析，我们得出结论，一些基因组重排是由于其功能重要性而保守的每个单元内序列之间的同源重组造成的。比较奈瑟氏球菌的4个基因组，我们鉴定出丝状噬菌体可能被其转座酶整合，并偶尔被其重排。4.限制性断裂等引发的DNA断裂和基因组重排我们发现，噬菌体单次感染后的III型限制性断裂可以通过原噬菌体的同源重组功能修复。研究发现同源驱动的非法重组依赖于 RecA 和 I 型限制位点。我们开发了一种灵敏的环状细菌染色体线性化测定方法，并分析了大肠杆菌的各种重组缺陷突变体。我们在大肠杆菌 BJ5183 菌株中展示了高水平的 DNA 双链断裂修复。我们模拟了Holliday结构的不对称分支迁移并分析了体外数据。5.限制性修饰基因复合物的扩增我们发现BamHI限制性修饰基因复合物在枯草芽孢杆菌染色体上的串联扩增依赖于限制性内切酶的功能。较少的