Investigation of chromosome dynamics in bacteria

与包含有丝分裂机制来分离染色体的真核细胞不同，原核生物如何将它们的DNA分离到未来的子细胞中尚不清楚。染色体凝聚和分离的关键因子是SMC蛋白。我们已经证明，对于真核细胞来说，SMC对于枯草杆菌的染色体分离是必不可少的。我们已经鉴定出两种蛋白质，SCPA和SCPB，它们在原核生物中保守，并与SMC形成三元复合体。荧光显微镜显示，SMC复合体以动态细胞周期依赖的方式定位，主要存在于靠近细胞极点的特定位置。更多的数据表明，SMC复合体从这些位置主动地将姐妹染色体拉向相反的细胞极。我们已经纯化了SMC的所有3种蛋白和不同的部分，结果表明，SCPA和SCPB结合到SMC的头域，而SMC以环状结构拥抱DNA，这呈现出一种新的DNA缩合机制，有待进一步阐明。我们还发现，DNA双链断裂修复所需的几种蛋白质在损伤诱导后动态和瞬时地招募到DNA上的特定位置，但以不同的方式进行。进一步的分析将提供对它们的功能和相互依存的详细洞察。

In contrast to eukaryotic cells that contain a mitotic machinery to separate chromosomes it is unclear how prokaryotes segregate their DNA into future daughter cells. Key players in chromosome condensation and segregation are SMC proteins. We have shown that likewise to eukaryotic cells SMC is essential for chromosome segregation in B. subtilis. We have identified two proteins, ScpA and ScpB that are conserved in prokaryotes and that form a ternary complex with SMC. Fluorescence microscopy has shown that the SMC complex localizes in a dynamic cell cycle dependent manner and is mostly present within specific sites close to the cell poles. Additional data suggest that from these sites the SMC complex actively pulls the sister chromosomes towards opposite cell poles. We have purified all 3 proteins and distinct parts of SMC and have shown that ScpA and ScpB bind to the SMC head domains, while SMC embraces DNA as a ring-like structure, which presents a novel DNA condensation mechanism that needs to be further elucidated. We have also found that several proteins required for repair of DNA double strand breaks are dynamically and transiently recruited to specific sites on the DNA after damage induction but in a differential fashion. Further analysis will provide detailed insight in their function and interdependence.