Genome-wide functional analysis reveals key roles for kinesins in the mammalian and mosquito stages of the malaria parasite life cycle

Abstract

Kinesins are microtubule-based motors important in cell division, motility, polarity, and intracellular transport in many eukaryotes, but poorly studied in eukaryotic pathogens. Plasmodium, the causative agent of malaria, is a divergent eukaryote with atypical aspects of cell division and plasticity of morphology throughout its lifecycle in both mammalian and mosquito hosts. Here we describe a genome-wide screen of all nine Plasmodium kinesins, revealing diverse subcellular locations and functions in spindle assembly, axoneme formation, and cell morphology. Surprisingly, only kinesin-13 has an essential role for growth in the mammalian host while the other eight kinesins are required during the proliferative and invasive stages of parasite transmission through the mosquito vector. In-depth analyses of kinesin-13 and kinesin-20 revealed functions in microtubule (MT) dynamics during apical polarity formation, spindle assembly, and axoneme biogenesis. This comprehensive study will inform the targeting of MT motors for therapeutic intervention in malaria.

Expansion microscopy of Plasmodium gametocytes reveals the molecular architecture of a bipartite microtubule organisation centre coordinating mitosis with axoneme assembly

“…Observations by electron and fluorescence microscopy gave general insights into the dynamics of cellular structures during microgametogenesis [4,[11][12][13] (S1 Fig) . The mature microgametocyte shows two electron-dense structures that have been linked with the formation of the mitotic spindles and the axonemes, respectively. One of this structure was described as an amorphous MTOC lying on the cytoplasmic face of a nuclear pore that is physically linked to another electron dense aggregation called the intranuclear body in the nuclear face of the same pore [9].…”

“…Our understanding of the ultrastructural organisation and cell division events of Plasmodium microgametocytes has initially relied on EM. It was more recently complemented by fluorescence live microscopy or super-resolution microscopy using multiple markers to infer the dynamic and the molecular composition of observed structures albeit at a lower resolution PLOS PATHOGENS [11][12][13]23,24]. Expansion microscopy (ExM) enables fluorophore labelling of proteins linked to a swellable polymer permitting isotropic physical expansion of the sample, which can be imaged using conventional microscopes below the diffraction limited resolution [25].…”

Mechanochemical tuning of a kinesin motor essential for malaria parasite transmission

“…Specifically, we used lysates of P. berghei gametocytes 6 min after activation because of the high expression of, and functional relevance for, the motor at this parasite life cycle stage [6][7][8] . Proteomic analysis of these samples identified a number of microtubule-associated proteins that are linked with male gamete maturation, axoneme formation and function, and are expressed specifically in male gametocytes, including dynein heavy chain 31 , kinesin-13 12 , calcium-dependent protein kinase 4 32 and PF16 33 (Supplementary Fig. 10a, b).…”

“…mammalian KIF18A, KIF18B, S. cerevisiae Kip3), kinesin-8Bs (mammalian KIF19A) and kinesin-8Xs 9 (Plasmodium kinesin-8X 11 ). Nine kinesins have been phylogenetically identified and were recently functionally characterised in P. berghei, 11,12 while at least 1 other may also be present in Plasmodium spp. genomes 6,8 .…”

Mechanochemical tuning of a kinesin motor essential for malaria parasite transmission

“…Future studies will reveal the cellular context in which these enzymes are utilised, leading to better understanding of divergence from canonical mammalian kinesins. Molecular understanding of these motors can thereby also contribute to potential new inhibition strategies for blocking key parasite life cycle transitions that rely on kinesins 53 .…”