Department of Microbiology and Molecular Genetics, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, Texas, USA.
Department of Microbiology and Molecular Genetics, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, Texas, USA.
J Biol Chem. 2022 Jun;298(6):101943. doi: 10.1016/j.jbc.2022.101943. Epub 2022 Apr 18.
Cytokinesis in the early divergent protozoan Trypanosoma brucei occurs from the anterior cell tip of the new-flagellum daughter toward the nascent posterior end of the old-flagellum daughter of a dividing biflagellated cell. The cleavage furrow ingresses unidirectionally along the preformed cell division fold and is regulated by an orphan kinesin named kinesin localized to the ingressing furrow (KLIF) that localizes to the leading edge of the ingressing furrow. Little is known about how furrow ingression is controlled by KLIF and whether KLIF interacts with and cooperates with other cytokinesis regulatory proteins to promote furrow ingression. Here, we investigated the roles of KLIF in cleavage furrow ingression and identified a cohort of KLIF-associated cytoskeletal proteins as essential cytokinesis regulators. By genetic complementation, we demonstrated the requirement of the kinesin motor activity, but not the putative tropomyosin domain, of KLIF in promoting furrow ingression. We further showed that depletion of KLIF impaired the resolution of the nascent posterior of the old-flagellar daughter cell, thereby stalking cleavage furrow ingression at late stages of cytokinesis. Through proximity biotinylation, we identified a subset of cytoskeleton-associated proteins (CAPs) as KLIF-proximal proteins, and functional characterization of these cytoskeletal proteins revealed the essential roles of CAP46 and CAP52 in positioning the cleavage furrow and the crucial roles of CAP42 and CAP50 in promoting cleavage furrow ingression. Together, these results identified multiple cytoskeletal proteins as cytokinesis regulators and uncovered their essential and distinct roles in cytokinesis.
在早期分化的原生动物布氏锥虫中，细胞分裂从新鞭毛子细胞的前端向旧鞭毛子细胞的新形成的后端进行。分裂双鞭毛细胞的分裂沟沿预先形成的细胞分裂褶皱单向内陷，由一种称为位于内陷沟的孤儿驱动蛋白（KLIF）的驱动蛋白调节，KLIF 定位于内陷沟的前缘。关于 KLIF 如何控制沟内陷以及 KLIF 是否与其他细胞分裂调节蛋白相互作用并合作以促进沟内陷知之甚少。在这里，我们研究了 KLIF 在分裂沟内陷中的作用，并确定了一组 KLIF 相关的细胞骨架蛋白是必不可少的细胞分裂调节蛋白。通过遗传互补，我们证明了 KLIF 促进沟内陷需要驱动蛋白的运动活性，但不需要假定的原肌球蛋白结构域。我们进一步表明，KLIF 的耗竭会损害旧鞭毛子细胞新形成的后端的分辨率，从而在细胞分裂的后期跟踪分裂沟内陷。通过邻近生物素化，我们确定了一组细胞骨架相关蛋白（CAPs）作为 KLIF 附近的蛋白，对这些细胞骨架蛋白的功能特征分析揭示了 CAP46 和 CAP52 在定位分裂沟以及 CAP42 和 CAP50 在促进分裂沟内陷中的关键作用。这些结果共同确定了多种细胞骨架蛋白作为细胞分裂调节蛋白，并揭示了它们在细胞分裂中的重要且不同的作用。
