Super-Resolution Microscopy and Single-Molecule Tracking Reveal Distinct Adaptive Dynamics of MreB and of Cell Wall-Synthesis Enzymes
Super-Resolution Microscopy and Single-Molecule Tracking Reveal Distinct Adaptive Dynamics of MreB and of Cell Wall-Synthesis Enzymes
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DOI：
10.3389/fmicb.2020.01946
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发表时间：
2020-08-20
影响因子：
5.2
通讯作者：
Graumann, Peter L.
中科院分区：
文献类型：
--
作者：
Dersch, Simon;Mehl, Johanna;Graumann, Peter L.
丝状、肌动蛋白样的MreB和合成细菌细胞壁的酶的运动被认为是高度协调的。我们研究了MreB、RodA和PbpH合成酶在500ms和20ms的时间尺度上的运动，使我们能够将整个MreB细丝以及单分子的运动与这两种合成蛋白质的运动进行比较。虽然这三种蛋白质都形成了以非常相似的轨迹方向和速度移动的组件，但它们的轨迹长度有很大差异，PbpH显示出最短的轨迹，MreB显示出最长的轨迹。这些实验表明，在假定的肽聚糖延伸机械(PGEM)以及在与MreB细丝相互作用期间，RodA和PbpH的开启/关闭速率不同。单分子跟踪显示PbpH和RodA存在明显的缓慢扩散和自由扩散的布居，表明它们在自由扩散和慢运动之间变化，表明它们与PGEM络合物是动态相互作用的。盐胁迫诱导后，MreB分子的动力学和微丝的取向和速度发生了显著变化，而RodA和PbpH单分子动力学变化不大。在胁迫适应阶段，细胞继续生长并延长细胞壁，而MreB形成的静止细丝越来越少。我们的结果表明，在胁迫适应过程中，细胞壁的合成是以MreB动力学适应的方式发生的，这表明枯草芽孢杆菌的细胞壁延伸涉及到具有不同结合动力学的酶与活性合成部位的相互作用。
The movement of filamentous, actin-like MreB and of enzymes synthesizing the bacterial cell wall has been proposed to be highly coordinated. We have investigated the motion of MreB and of RodA and PbpH cell wall synthesis enzymes at 500 ms and at 20 ms time scales, allowing us to compare the motion of entire MreB filaments as well as of single molecules with that of the two synthesis proteins. While all three proteins formed assemblies that move with very similar trajectory orientation and with similar velocities, their trajectory lengths differed considerably, with PbpH showing shortest and MreB longest trajectories. These experiments suggest different on/off rates for RodA and PbpH at the putative peptidoglycan-extending machinery (PGEM), and during interaction with MreB filaments. Single molecule tracking revealed distinct slow-moving and freely diffusing populations of PbpH and RodA, indicating that they change between free diffusion and slow motion, indicating a dynamic interaction with the PGEM complex. Dynamics of MreB molecules and the orientation and speed of filaments changed markedly after induction of salt stress, while there was little change for RodA and PbpH single molecule dynamics. During the stress adaptation phase, cells continued to grow and extended the cell wall, while MreB formed fewer and more static filaments. Our results show that cell wall synthesis during stress adaptation occurs in a mode involving adaptation of MreB dynamics, and indicate thatBacillus subtiliscell wall extension involves an interplay of enzymes with distinct binding kinetics to sites of active synthesis.