Functional Characterization of the Role of the Chromosome I Partitioning System in Genome Segregation in Deinococcus radiodurans
Functional Characterization of the Role of the Chromosome I Partitioning System in Genome Segregation in Deinococcus radiodurans
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DOI：
10.1128/jb.00610-12
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发表时间：
2012-11-01
影响因子：
3.2
通讯作者：
Misra, Hari S.
中科院分区：
文献类型：
--
作者：
Charaka, Vijay Kumar;Misra, Hari S.
耐辐射异常球菌是一种耐辐射细菌，具有多组分基因组。染色体I含有三个推定的着丝粒（segS 1、segS 2和segS 3），以及帕拉（ParA 1）和帕拉B（ParB 1）同源物。ParB 1与segS的相互作用是序列特异性的，ParA 1被证明是DNA结合ATP酶。ParA 1的ATP酶活性的刺激时，segS元素与ParB 1共孵育，但最大的增加与segS 3观察。ParA 1与segS-ParB 1复合物一起孵育，在不存在ATP的情况下显示出增加的光散射。在ATP的存在下，这种增加继续与segS 1-ParA 1B 1和segS 2-ParA 1B 1复合物，而它在segS 3的情况下，在初始增加30分钟后迅速下降。D.表达绿色荧光蛋白（GFP）-ParB 1的耐辐射细胞在类核上产生病灶，并且Δ parB 1突变体显示出生长迟缓，并且与野生型相比，无核化高出13%。携带segS 1和segS 2的不稳定的mini-F质粒在没有ParA 1B 1的大肠杆菌中表现出遗传性，而segS 3介导的质粒稳定性需要ParA 1B 1的反式表达。与未转化的E.在大肠杆菌细胞中，携带pDAGS 3（携带segS 3的质粒并且还表达ParB 1-GFP）的细胞在类核上产生离散的GFP焦点。这些结果表明，D.抗辐射蛋白具有功能活性，并在基因组分离中起作用。
Deinococcus radiodurans, a radiation-resistant bacterium, harbors a multipartite genome. Chromosome I contains three putative centromeres (segS1, segS2, and segS3), and ParA (ParA1) and ParB (ParB1) homologues. The ParB1 interaction with segS was sequence specific, and ParA1 was shown to be a DNA binding ATPase. The ATPase activity of ParA1 was stimulated when segS elements were coincubated with ParB1, but the greatest increase was observed with segS3. ParA1 incubated with the segS-ParB1 complex showed increased light scattering in the absence of ATP. In the presence of ATP, this increase was continued with segS1-ParA1B1 and segS2-ParA1B1 complexes, while it decreased rapidly after an initial increase for 30 min in the case of segS3. D. radiodurans cells expressing green fluorescent protein (GFP)-ParB1 produced foci on nucleoids, and the Delta parB1 mutant showed growth retardation and similar to 13%-higher anucleation than the wild type. Unstable mini-F plasmids carrying segS1 and segS2 showed inheritance in Escherichia coli without ParA1B1, while segS3-mediated plasmid stability required the in trans expression of ParA1B1. Unlike untransformed E. coli cells, cells harboring pDAGS3, a plasmid carrying segS3 and also expressing ParB1-GFP, produced discrete GFP foci on nucleoids. These findings suggested that both segS elements and the ParA1B1 proteins of D. radiodurans are functionally active and have a role in genome segregation.