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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.我们发现EcoRI修饰酶与EcoRI限制性内切酶和大量细胞蛋白一样稳定。我们用进化生态学的方法分析了成瘾基因的进化条件。论证了空间结构的重要性。通过对深渊焦球菌（Pyrococcus abyssi）和博氏焦球菌（Pyrococcus horikoshii）基因组的比较，我们发现了一个插入到前者基因组中的限制性修饰基因复合物。通过筛选小麦胚源性无细胞蛋白合成系统中的限制性内切酶pai。它在5‘GTAC.3处产生一个新的末端TA3’。通过比较近缘基因组重建大基因组多态性的形成通过比较7个金黄色葡萄球菌的基因组，我们发现I型限制性内切酶的S（特异性）亚基通过两个目标识别结构域的结合而进化。通过对三个串联同源基因簇的分析，我们得出结论，一些基因组重排是由于其功能重要性保守的每个单元内序列之间的同源重组造成的。比较奈瑟菌的4个基因组，我们发现丝状噬菌体可能被它的转座酶整合，偶尔也会被它重新排列。我们发现，单次感染噬菌体lambda后，III型限制性内切酶断裂可通过噬菌体的同源重组功能进行修复。发现同源驱动的非法重组依赖于RecA和I型限制位点。我们建立了一种灵敏的环形细菌染色体线性化试验，并分析了大肠杆菌的各种重组缺陷突变体。结果表明，大肠杆菌BJ5183具有高水平的DNA双链断裂修复能力。建立了Holliday结构的非对称分支迁移模型，并对数据进行了体外分析。限制性修饰基因复合物的增殖研究表明，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。少