Model-based trajectory classification of anchored molecular motor-biopolymer interactions.
Model-based trajectory classification of anchored molecular motor-biopolymer interactions.
复制标题
DOI：
10.1016/j.bpr.2023.100130
复制
发表时间：
2023-12-13
期刊：
影响因子：
--
通讯作者：
中科院分区：
文献类型：
--
作者：
在许多物种的合子有丝分裂过程中，细胞皮层产生的力是父系提供的中心体分离和迁移、原核迁移、遗传物质分离和细胞分裂所必需的。此外，在某些物种中，纺锤体微管和皮质之间的力产生相互作用将有丝分裂纺锤体不对称地定位在受精卵内，这是不对称细胞分裂的重要步骤。理解微管依赖的力产生的机械和分子机制，因此不对称细胞分裂需要识别个体皮质力产生单位在体内。目前还没有一种方法能够以高的时空分辨率来识别单个的力生成单元。在这里，我们提出了一种方法来确定的位置和相对数量的微管依赖性皮质力产生单位使用荧光标记的动力蛋白的单分子成像。动力蛋白的行为进行建模，分类皮质结合动力蛋白的轨迹，根据他们是否与微管相互作用。分类策略概括了秀丽隐杆线虫受精卵有丝分裂中众所周知的力不对称性。为了评估分类的稳健性，我们使用RNAi来耗尽微管蛋白亚基TBA-2。正如预测的那样，这种处理减少了被归类为与微管接合的轨迹的数量。我们的技术将是一个有价值的工具，以定义动力蛋白皮质力产生及其调节的分子机制，以及锚定马达与生物聚合物相互作用的其他情况（例如，肌动蛋白、微管蛋白、DNA）。
During zygotic mitosis in many species, forces generated at the cell cortex are required for the separation and migration of paternally provided centrosomes, pronuclear migration, segregation of genetic material, and cell division. Furthermore, in some species, force-generating interactions between spindle microtubules and the cortex position the mitotic spindle asymmetrically within the zygote, an essential step in asymmetric cell division. Understanding the mechanical and molecular mechanisms of microtubule-dependent force generation and therefore asymmetric cell division requires identification of individual cortical force-generating units in vivo. There is no current method for identifying individual force-generating units with high spatiotemporal resolution. Here, we present a method to determine both the location and the relative number of microtubule-dependent cortical force-generating units using single-molecule imaging of fluorescently labeled dynein. Dynein behavior is modeled to classify trajectories of cortically bound dynein according to whether they are interacting with a microtubule. The categorization strategy recapitulates well-known force asymmetries in C. elegans zygote mitosis. To evaluate the robustness of categorization, we used RNAi to deplete the tubulin subunit TBA-2. As predicted, this treatment reduced the number of trajectories categorized as engaged with a microtubule. Our technique will be a valuable tool to define the molecular mechanisms of dynein cortical force generation and its regulation as well as other instances wherein anchored motors interact with biopolymers (e.g., actin, tubulin, DNA).