Research Article: An orphan kinesin controls trypanosome morphology transitions by targeting FLAM3 to the flagellum

Date Published: May 29, 2018

Publisher: Public Library of Science

Author(s): Tai An, Ziyin Li, Kent L. Hill.


Trypanosoma brucei undergoes life cycle form transitions from trypomastigotes to epimastigotes in the insect vector by re-positioning the mitochondrial genome and re-locating the flagellum and flagellum-associated cytoskeletal structures. The mechanism underlying these dramatic morphology transitions remains poorly understood. Here we report the regulatory role of the orphan kinesin KIN-E in controlling trypanosome morphology transitions. KIN-E localizes to the flagellum and is enriched at the flagellar tip, and this localization depends on the C-terminal m-calpain domain III-like domains. Depletion of KIN-E in the trypomastigote form of T. brucei causes major morphology changes and a gradual increase in the level of EP procyclin, generating epimastigote-like cells. Mechanistically, through its C-terminal importin α-like domain, KIN-E targets FLAM3, a flagellar protein involved in morphology transitions, to the flagellum to promote elongation of the flagellum attachment zone and positioning of the flagellum and flagellum-associated cytoskeletal structure, thereby maintaining trypomastigote cell morphology. Our findings suggest that morphology transitions in trypanosomes require KIN-E-mediated transport of FLAM3 to the flagellum.

Partial Text

Trypanosomatids, a group of protozoan parasites consisting of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania spp., transition to different developmental forms with distinct cell morphology during their life cycle between the insect vectors and the mammalian hosts. These life cycle forms are distinguished by the relative position of the kinetoplast, the cell’s mitochondrial genome, to the nucleus, the cellular location from which the flagellum emerges, and the length of the free, unattached flagellum [1]. Inside the proventriculus of the tsetse fly vector, T. brucei differentiates from trypomastigote form to epimastigote form, which then undergoes an asymmetrical cell division and further develops to metacyclic form, the mammal-infective form of the parasite, in the salivary gland [1]. Although the molecular mechanisms underlying the transitions between these life cycle forms in trypanosomatids remain poorly understood, several proteins, including some RNA-binding proteins and a few flagellum-associated cytoskeletal proteins, were recently found to be involved in life cycle transitions in T. brucei [2,3,4,5,6,7]. The involvement of RNA-binding proteins ALBA3/4 [3] and RBP6 [2] in trypanosome life cycle transitions suggests a posttranscriptional regulation scheme, but mechanistically how these proteins contribute to this process is still elusive. The involvement of two flagellum attachment zone (FAZ) proteins in the flagellum, ClpGM6 and FLAM3 [4,5], and two intracellular FAZ proteins, FAZ9 [6] and TbSAS-4 [7], in life cycle form transitions suggests that the morphology transitions require the modulation of flagellum-associated cytoskeletal structures mediated by these FAZ proteins.

Morphology transitions during trypanosome life cycle development appear to involve the modulation of the length of the FAZ through a cohort of FAZ flagellum domain proteins, such as ClpGM6 [5] and FLAM3 [4,14], and some intracellular FAZ proteins, such as FAZ9 [6] and TbSAS-4 [7]. Here an orphan kinesin, KIN-E, was found to play an essential role in controlling morphology transitions in T. brucei (Fig 3). KIN-E is unusual in that it contains in its C-terminus an importin α-like domain and two m-calpain domain III-like domains (Fig 1), which have not been found in any other kinesins in any organisms. The importin α-like domain and the m-calpain domain III-like domains play distinct roles in regulating KIN-E function. The importin α-like domain is not required for KIN-E localization (Fig 6A), but it is essential for KIN-E function (Fig 6C) and for interacting with FLAM3 and transporting the latter to the flagellum (Fig 8C). The m-calpain domain III-like domains are not involved in binding to FLAM3 (Fig 8A and 8B), but are essential for KIN-E localization to the flagellum (Fig 8C) and for KIN-E function (Fig 6C). The domain III in calpain, a calcium-dependent cysteine protease in vertebrates, functions as a calcium-regulated phospholipid-binding domain [22]. Thus, the two m-calpain domain III-like domains in KIN-E may also be capable of binding to calcium and lipid. The biochemical function and the mechanistic role of the m-calpain domain III-like domains in mediating KIN-E localization require further investigation.