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Cytokinesis in Trypanosoma brucei relies on an orphan kinesin that dynamically crosslinks microtubules.

ABSTRACT: Many single-celled eukaryotes have complex cell morphologies defined by microtubules arranged into higher-order structures. The auger-like shape of the parasitic protist Trypanosoma brucei (T. brucei) is mediated by a parallel array of microtubules that underlies the plasma membrane. The subpellicular array must be partitioned and segregated using a microtubule-based mechanism during cell division. We previously identified an orphan kinesin, KLIF, that localizes to the ingressing cleavage furrow and is essential for the completion of cytokinesis. We have characterized the biophysical properties of a truncated KLIF construct in vitro to gain mechanistic insight into the function of this novel kinesin. We find that KLIF is a non-processive dimeric kinesin that dynamically crosslinks microtubules. Microtubules crosslinked by KLIF in an antiparallel orientation are translocated relative to one another, while microtubules crosslinked parallel to one another remain static, resulting in the formation of organized parallel bundles. In addition, we find that KLIF stabilizes the alignment of microtubule plus ends. These features provide a mechanistic understanding for how KLIF functions to form a new pole of aligned microtubule plus ends that defines the shape of the new cell posterior, which is an essential requirement for the completion of cytokinesis in T. brucei.

SUBMITTER: Sladewski TE

PROVIDER: S-EPMC10023446 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Cytokinesis in Trypanosoma brucei relies on an orphan kinesin that dynamically crosslinks microtubules.

Sladewski Thomas E TE Campbell Paul C PC Billington Neil N D'Ordine Alexandra A Cole James L JL de Graffenried Christopher L CL

Current biology : CB 20230213 5

Many single-celled eukaryotes have complex cell morphologies defined by microtubules arranged into higher-order structures. The auger-like shape of the parasitic protist Trypanosoma brucei (T. brucei) is mediated by a parallel array of microtubules that underlies the plasma membrane. The subpellicular array must be partitioned and segregated using a microtubule-based mechanism during cell division. We previously identified an orphan kinesin, KLIF, that localizes to the ingressing cleavage furrow ...[more]

PMID: 36787745

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Project description:Kinesins are microtubule-based motor proteins that play diverse cellular functions by regulating microtubule dynamics and intracellular transport in eukaryotes. The early branching kinetoplastid protozoan Trypanosoma brucei has an expanded repertoire of kinetoplastid-specific kinesins and orphan kinesins, many of which have unknown functions. We report here the identification of an orphan kinesin named KIN-G that plays an essential role in maintaining hook complex integrity and promoting Golgi biogenesis in T. brucei. KIN-G localizes to the distal portion of the centrin arm of the flagellum-associated hook complex through association with the centrin arm protein TbCentrin4. Knockdown of KIN-G in T. brucei disrupts the integrity of the hook complex by reducing the length of the centrin arm and eliminating the shank part of the hook complex, thereby impairing flagellum attachment zone elongation and flagellum positioning, which leads to unequal cytokinesis. KIN-G associates with Golgi through a centrin arm-localized Golgi peripheral protein named CAAP1, which maintains Golgi-centrin arm association to facilitate Golgi biogenesis. Knockdown of KIN-G impairs Golgi biogenesis by disrupting CAAP1 at the centrin arm, thereby impairing the maturation of centrin arm-associated Golgi. In vitro microtubule gliding assays demonstrate that KIN-G is a plus end-directed motor protein, and its motor activity is required for hook complex assembly and Golgi biogenesis. Together, these results identify a kinesin motor protein for promoting hook complex assembly and uncover a control mechanism for Golgi biogenesis through KIN-G-mediated maintenance of Golgi-hook complex association.IMPORTANCETrypanosoma brucei has a motile flagellum, which controls cell motility, cell morphogenesis, cell division, and cell-cell communication, and a set of cytoskeletal structures, including the hook complex and the centrin arm, associates with the flagellum. Despite the essentiality of these flagellum-associated cytoskeletal structures, their mechanistic roles and the function of their associated proteins remain poorly understood. Here, we demonstrate that the orphan kinesin KIN-G functions to promote the biogenesis of the hook complex and the Golgi apparatus. KIN-G exerts this function by mediating the association between centrin arm and Golgi through the centrin arm protein TbCentrin4 and a novel Golgi scaffold protein named CAAP1, thereby bridging the two structures and maintaining their close association to facilitate the assembly of the two structures. These findings uncover the essential involvement of a kinesin motor protein in regulating the biogenesis of the hook complex and the Golgi in trypanosomes.

Dataset Information

Cytokinesis in Trypanosoma brucei relies on an orphan kinesin that dynamically crosslinks microtubules.

Publications

Cytokinesis in Trypanosoma brucei relies on an orphan kinesin that dynamically crosslinks microtubules.

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