Duellberg Christian, Cade Nicholas I, Holmes David, Surrey Thomas
Lincoln's Inn Fields Laboratory, The Francis Crick Institute, London, United Kingdom.
London Centre of Nanotechnology, London, United Kingdom.
Elife. 2016 Apr 6;5:e13470. doi: 10.7554/eLife.13470.
The function of microtubules relies on their ability to switch between phases of growth and shrinkage. A nucleotide-dependent stabilising cap at microtubule ends is thought to be lost before this switch can occur; however, the nature and size of this protective cap are unknown. Using a microfluidics-assisted multi-colour TIRF microscopy assay with close-to-nm and sub-second precision, we measured the sizes of the stabilizing cap of individual microtubules. We find that the protective caps are formed by the extended binding regions of EB proteins. Cap lengths vary considerably and longer caps are more stable. Nevertheless, the trigger of instability lies in a short region at the end of the cap, as a quantitative model of cap stability demonstrates. Our study establishes the spatial and kinetic characteristics of the protective cap and provides an insight into the molecular mechanism by which its loss leads to the switch from microtubule growth to shrinkage.
微管的功能依赖于它们在生长和收缩阶段之间切换的能力。在这种切换发生之前，微管末端依赖核苷酸的稳定帽被认为会丢失；然而，这种保护帽的性质和大小尚不清楚。我们使用具有接近纳米和亚秒级精度的微流控辅助多色全内反射荧光显微镜测定法，测量了单个微管稳定帽的大小。我们发现保护帽是由EB蛋白的延伸结合区域形成的。帽的长度差异很大，较长的帽更稳定。然而，正如帽稳定性的定量模型所示，不稳定性的触发因素在于帽末端的一个短区域。我们的研究确定了保护帽的空间和动力学特征，并深入了解了其丢失导致微管从生长转变为收缩的分子机制。
