Imaging DivIVA dynamics using photo-convertible and activatable fluorophores in Bacillus subtilis
Imaging DivIVA dynamics using photo-convertible and activatable fluorophores in Bacillus subtilis
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DOI：
10.3389/fmicb.2014.00059
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发表时间：
2014-02-18
影响因子：
5.2
通讯作者：
Bramkamp, Marc
中科院分区：
文献类型：
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作者：
Bach, Juri N.;Albrecht, Nadine;Bramkamp, Marc
大多数杆状模型生物体，例如大肠杆菌或枯草芽孢杆菌，利用两种抑制系统来正确定位细胞分裂装置。虽然类核封闭系统在类核附近起作用，但 Min 系统被认为可以保护细胞极免于徒劳分裂，从而导致无 DNA 的微型细胞。 Min 系统由抑制蛋白 MinC 组成，它在 FtsZ 环形成水平上发挥作用。 MinC 被 MinD 招募到膜上，MinD 是 Walker-ATP 酶 MinD/ParA 家族的成员。 MinCD 复合体的拓扑定位取决于大肠杆菌中的 MinE 和枯草芽孢杆菌中的 MinJ/DivIVA。虽然 MinE 在大肠杆菌细胞中驱动 MinCD 振荡，在细胞中部具有时间依赖性最小浓度，但枯草芽孢杆菌系统被认为通过 MinJ/DivIVA 稳定地束缚在细胞极上。最近的发展表明，枯草芽孢杆菌中的 Min 系统主要在分裂部位发挥作用，在那里它似乎阻止了分裂机制的重新启动。因此，MinCD 描述了枯草芽孢杆菌的动态行为。这与 DivIVA 在细胞两极的稳定定位有些不一致。正在进行的分裂的高分辨率成像显示 DivIVA 也在分裂部位富集。在这里，我们分析极局域 DivIVA 是否具有部分移动性并且可以对隔膜 DivIVA 产生影响，反之亦然。为此，我们使用绿色至红色光转换荧光团、Dendra2 和光激活 PA-GFP 的融合。事实证明，这些技术对于区分体内蛋白质重新定位非常有效。我们的结果表明，枯草芽孢杆菌 DivIVA 确实是动态的，并且从两极移动到新的隔膜。
Most rod-shape model organisms such as Escherichia coli or Bacillus subtilis utilize two inhibitory systems for correct positioning of the cell division apparatus. While the nucleoid occlusion system acts in vicinity of the nucleoid, the Min system was thought to protect the cell poles from futile division leading to DNA-free miniature cells. The Min system is composed of an inhibitory protein, MinC, which acts at the level of the FtsZ ring formation. MinC is recruited to the membrane by MinD, a member of the MinD/ParA family of Walker-ATPases. Topological positioning of the MinCD complex depends on MinE in E. coli and MinJ/DivIVA in B. subtilis. While MinE drives an oscillation of MinCD in the E. coli cell with a time-dependent minimal concentration at midcell, the B. subtilis system was thought to be stably tethered to the cell poles by MinJ/DivIVA. Recent developments revealed that the Min system in B. subtilis mainly acts at the site of division, where it seems to prevent reinitiation of the division machinery. Thus, MinCD describe a dynamic behavior in B. subtilis. This is somewhat inconsistent with a stable localization of DivIVA at the cell poles. High resolution imaging of ongoing divisions show that DivIVA also enriches at the site of division. Here we analyze whether polar localized DivIVA is partially mobile and can contribute to septal DivIVA and vice versa. For this purpose we use fusions with green to red photoconvertible fluorophores, Dendra2 and photoactivatable PA-GFP. These techniques have proven very powerful to discriminate protein relocalization in vivo. Our results show that B. subtilis DivIVA is indeed dynamic and moves from the poles to the new septum.