Distribution of stable DnaA-binding sites on the Bacillus subtilis genome detected using a modified ChIP-chip method
Distribution of stable DnaA-binding sites on the Bacillus subtilis genome detected using a modified ChIP-chip method
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DOI：
10.1093/dnares/dsm017
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发表时间：
2007-08-31
期刊：
影响因子：
4.1
通讯作者：
Ogasawara, Naotake
中科院分区：
文献类型：
--
作者：
Ishikawa, Shu;Ogura, Yoshitoshi;Ogasawara, Naotake
我们开发了一种改进的ChIP芯片方法，命名为ChAP芯片（染色质亲和沉淀与平铺芯片耦合）。利用该技术测定的枯草芽孢杆菌Spo 0 J的结合位点与以前的研究结果一致。DNA复制起始蛋白DnaA在B上的8个基因间区域形成稳定的复合物。枯草杆菌基因组结合序列的表征表明，两个因素的DnaA盒的局部密度和它们对DnaA的亲和力是稳定结合的关键。我们进一步表明，除了自身调节外，DnaA直接正向调节sda的表达，而ywlC和ywlA以负向方式调节。在其他芽孢杆菌属物种中可能的稳定DNA结合序列的检查表明，DNA依赖性调节这些基因保持在大多数细菌检查，支持其生物学意义。此外，一个可能的稳定的DnaA结合位点下游的gcp也建议是保守的。此外，已经鉴定了对每种细菌特异性的潜在DnaA结合序列，通常与oriC非常接近。这些研究结果表明，DnaA发挥了几个额外的作用，如控制有效的启动子，ATP-DnaA的水平，和/或稳定的oriC周围的基因组结构域结构的染色体复制的正确启动。
We developed a modified ChIP-chip method, designated ChAP-chip (Chromatin Affinity Precipitation coupled with tiling chip). The binding sites of Bacillus subtilis Spo0J determined using this technique were consistent with previous findings. A DNA replication initiator protein, DnaA, formed stable complexes at eight intergenic regions on the B. subtilis genome. Characterization of the binding sequences suggested that two factors-the local density of DnaA boxes and their affinities for DnaA-are critical for stable binding. We further showed that in addition to autoregulation, DnaA directly modulate the expression of sda in a positive, and ywlC and yydA in a negative manner. Examination of possible stable DnaA-binding sequences in other Bacillus species suggested that DnaA-dependent regulation of those genes is maintained in most bacteria examined, supporting their biological significance. In addition, a possible stable DnaA-binding site downstream of gcp is also suggested to be conserved. Furthermore, potential DnaA-binding sequences specific for each bacterium have been identified, generally in close proximity to oriC. These findings suggest that DnaA plays several additional roles, such as control of the level of effective initiator, ATP-DnaA, and/or stabilization of the domain structure of the genome around oriC for the proper initiation of chromosome replication.