Conjugative DNA Transfer Is Enhanced by Plasmid R1 Partitioning Proteins.
Conjugative DNA Transfer Is Enhanced by Plasmid R1 Partitioning Proteins.
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DOI：
10.3389/fmolb.2016.00032
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发表时间：
2016
影响因子：
5
通讯作者：
Zechner EL
中科院分区：
文献类型：
--
作者：
Gruber CJ;Lang S;Rajendra VK;Nuk M;Raffl S;Schildbach JF;Zechner EL
细菌接合是IV型分泌的一种形式，用于将蛋白质和DNA直接转运到受体细菌。该过程是细胞接触依赖性的，然而使细胞外事件触发质粒转移到细胞内开始的机制仍然不清楚。在本研究中，我们研究了质粒R1蛋白在基因转移起始中的作用。我们发现，TraI，接合DNA加工的中央调节器，相互作用的物理和功能与质粒分配蛋白ParM和ParR。这些相互作用在体内和体外刺激TraI催化的质粒DNA松弛，并增加ParM ATP酶活性。ParM还结合偶联蛋白TraD和VirB 4样通道ATP酶TraC。总之，这些蛋白质-蛋白质相互作用可能起到细胞内共定位转移组分并促进缀合机器的组装的作用。重要的是，这些数据还表明，ParM和ParR在接合孔处的持续缔合对于质粒转移有效地开始是必需的。此外，在不存在Par蛋白的情况下组装的接合菌毛和潜在的分泌机制介导较差的生物膜形成，并且对于菌毛特异性R17噬菌体摄取完全功能失调。因此，在松弛体，偶联蛋白和通道ATP酶的界面上的Par组件的功能整合对于最佳构象和有效激活转移机器显得重要。我们的结论是，低拷贝质粒R1已经演变成一个积极的隔离系统，优化其垂直和横向传播模式。
Bacterial conjugation is a form of type IV secretion used to transport protein and DNA directly to recipient bacteria. The process is cell contact-dependent, yet the mechanisms enabling extracellular events to trigger plasmid transfer to begin inside the cell remain obscure. In this study of plasmid R1 we investigated the role of plasmid proteins in the initiation of gene transfer. We find that TraI, the central regulator of conjugative DNA processing, interacts physically, and functionally with the plasmid partitioning proteins ParM and ParR. These interactions stimulate TraI catalyzed relaxation of plasmid DNA in vivo and in vitro and increase ParM ATPase activity. ParM also binds the coupling protein TraD and VirB4-like channel ATPase TraC. Together, these protein-protein interactions probably act to co-localize the transfer components intracellularly and promote assembly of the conjugation machinery. Importantly these data also indicate that the continued association of ParM and ParR at the conjugative pore is necessary for plasmid transfer to start efficiently. Moreover, the conjugative pilus and underlying secretion machinery assembled in the absence of Par proteins mediate poor biofilm formation and are completely dysfunctional for pilus specific R17 bacteriophage uptake. Thus, functional integration of Par components at the interface of relaxosome, coupling protein, and channel ATPases appears important for an optimal conformation and effective activation of the transfer machinery. We conclude that low copy plasmid R1 has evolved an active segregation system that optimizes both its vertical and lateral modes of dissemination.