Abstract
Kinesin-5 motors play essential roles in spindle apparatus assembly during cell division, by generating forces to establish and maintain the spindle bipolarity essential for proper chromosome segregation. Kinesin-5 is largely conserved structurally and functionally in model eukaryotes, but its role is unknown in the Plasmodium parasite, an evolutionarily divergent organism with several atypical features of both mitotic and meiotic cell division. We have investigated the function and subcellular location of kinesin-5 during cell division throughout the Plasmodium berghei life cycle. Deletion of kinesin-5 had little visible effect at any proliferative stage except sporozoite production in oocysts, resulting in a significant decrease in the number of motile sporozoites in mosquito salivary glands, which were able to infect a new vertebrate host. Live-cell imaging showed kinesin-5-GFP located on the spindle and at spindle poles during both atypical mitosis and meiosis. Fixed-cell immunofluorescence assays revealed kinesin-5 co-localized with α-tubulin and centrin-2 and a partial overlap with kinetochore marker NDC80 during early blood stage schizogony. Dual-color live-cell imaging showed that kinesin-5 is closely associated with NDC80 during male gametogony, but not with kinesin-8B, a marker of the basal body and axonemes of the forming flagella. Treatment of gametocytes with microtubule-specific inhibitors confirmed kinesin-5 association with nuclear spindles and not cytoplasmic axonemal microtubules. Altogether, our results demonstrate that kinesin-5 is associated with the spindle apparatus, expressed in proliferating parasite stages, and important for efficient production of infectious sporozoites.
Highlights
Kinesin-5 proteins are a family of molecular motors that is structurally and functionally conserved throughout eukaryotes (Wojcik et al, 2013; Waitzman and Rice, 2014; Mann and Wadsworth, 2019)
Spindle apparatus assembly and chromosome segregation are key processes of nuclear division that require forces generated by MT-based motor proteins (Wordeman, 2010; Kull and Endow, 2013)
In the present study we show by deletion of kinesin-5 that this protein is not essential for either mitotic or meiotic division during P. berghei parasite proliferation but it has an important role in production of infectious sporozoites
Summary
Kinesin-5 proteins are a family of molecular motors that is structurally and functionally conserved throughout eukaryotes (Wojcik et al, 2013; Waitzman and Rice, 2014; Mann and Wadsworth, 2019). They are involved in spindle pole separation and are considered essential for mitosis in the vast majority of eukaryotes (Bannigan et al, 2007; Ferenz et al, 2010), except Caenorhabditis elegans (Bishop et al, 2005), Dictyostelium discoideum (Tikhonenko et al, 2008), and Candida albicans (Shoukat et al, 2019). The kinesin-5 motor domain is conserved across eukaryotes including Plasmodium (Supplementary Figure 1A), but the tail region is highly variable except for a short, conserved region called the BimC box, which contains consensus sites for phosphorylation by cyclin dependent kinase 1 (Cdk1) in most eukaryotes (Sharp et al, 1999; Bishop et al, 2005; Chee and Haase, 2010)
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