<i>Plasmodium</i>centrin<i>Pb</i>CEN-4 localizes to the putative MTOC and is dispensable for malaria parasite proliferation

Roques, Magali; Stanway, Rebecca R.; Rea, Edward; Márkus, Róbert; Brady, Declan; Holder, Anthony A.; Guttery, David S.; Tewari, Rita

doi:10.1242/bio.036822

Cited by 33 publications

(56 citation statements)

References 40 publications

(53 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The asynchronous nature of the division during these stages is shown by nuclei having both one and two NDC80-GFP foci (and intermediate forms) within the same cell. We made similar observations in our previous study of CEN-4 (Roques et al, 2019). These studies also suggest that the kinetochore and centrosome/spindle pole body duplicate before nuclear division starts; therefore the duplication of NDC80 and CEN-4 sets the stage for mitosis at each round of nuclear division in Plasmodium, as in Toxoplasma (Suvorova et al, 2015).…”

Section: Discussionsupporting

confidence: 88%

“…Live cell imaging of transgenic parasite lines was performed at different proliferative stages during the parasite life cycle (Figure 1) as described previously (Roques et al, 2019;Saini et al, 2017) using a Zeiss AxioImager M2 microscope fitted with an AxioCam ICc1 digital camera (Carl Zeiss, Inc).…”

Section: Localization Of Ndc80-gfp Throughout the Parasite Life Cyclementioning

confidence: 99%

“…The process of chromosome segregation and kinetochore dynamics throughout the life cycle are not well understood. To date, analysis of Plasmodium mitotic/meiotic spindle assembly and chromosome segregation has been performed largely using transmission electron microscopy (TEM) (Sinden et al, 1978;Sinden et al, 1976), and biochemical analysis of microtubule markers including α -tubulin (Fennell et al, 2008), and centrins associated with the putative MTOC (Gerald et al, 2011;Roques et al, 2019). An analysis of a Plasmodium artificial chromosome (PAC) identified a putative centrosome derived from chromosome 5 (PbCEN5), and highlighted the dynamics of chromosome segregation during both mitotic and meiotic stages of the parasite's life cycle (Iwanaga et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Real-time dynamics of Plasmodium NDC80 reveals unusual modes of chromosome segregation during parasite proliferation

Pandey

Zeeshan

Ferguson

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

Eukaryotic cell proliferation requires chromosome replication and precise segregation, followed by cell division, to ensure daughter cells have identical genomic copies. The kinetochore is a multi-protein structure assembled on chromosome centromeres, which mediates attachment to spindle microtubules and chromosome segregation during mitosis. Plasmodium spp., the causative agent of malaria, undergoes closed mitosis with an intact nuclear membrane and intranuclear mitotic spindle. However, the regulation of the main events of mitosis and meiosis, including chromosome segregation, is poorly understood in this parasite. In part this is due to a distinct lack of markers for tracking kinetochore dynamics. Since NDC80 is the conserved component of the kinetochore complex, we labelled NDC80 with either a C-terminal GFP or mCherry tag by recombination at the endogenous locus to generate transgenic lines of the rodent malaria parasite Plasmodium berghei. We show that NDC80 is located at a single cluster of all kinetochores and examine its spatio-temporal dynamics throughout all proliferative stages of the parasite life cycle.We also compare the location of NDC80 with that of the kinetochore using electron microscopy. These transgenic parasite lines are a very useful resource to identify and study the kinetochore complex and follow chromosome dynamics during malaria parasite development.

show abstract

Section: Discussionsupporting

confidence: 88%

Section: Localization Of Ndc80-gfp Throughout the Parasite Life Cyclementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Real-time dynamics of Plasmodium NDC80 reveals unusual modes of chromosome segregation during parasite proliferation

Pandey

Zeeshan

Ferguson

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…Kinesin-5-GFP fluorescence was observed as multiple foci representing a location at the putative MTOC/nuclear poles, from very early in development (day 7) to late stages (at day 14) of oocyst maturation (Fig. 6B), similar to the pattern for PbCEN-4 as described previously [36].…”

Section: Pbkinesin-5-gfp Exhibits Multiple Nuclear Foci During Oocystsupporting

confidence: 83%

Plasmodium bergheikinesin-5 associates with the spindle apparatus during cell division and is important for efficient production of infectious sporozoites

Zeeshan

Brady

Stanway

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

AbstractKinesin-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. The only significant effect was a 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 exhibited a partial overlap with kinetochore marker NDC80 during early blood stage schizogony. Dual-colour live-cell imaging during male gametogony showed that kinesin-5 is closely associated with NDC80, 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 and expressed in proliferating parasite stages, but it is not essential for parasite survival.ImportanceKinesin-5 is a motor protein with important roles during cell proliferation and therefore considered a strong target for therapeutics development against many diseases. The role of kinesin-5 in Plasmodium is unknown, but in this organism there are atypical aspects of cell division that differ from those of model eukaryotes. For example, classical regulators such as polo-like kinase, classical cyclins and cyclin dependent kinases are absent from Plasmodium. Here, we show that in Plasmodium, unlike in most model eukaryotes, kinesin-5 is dispensable, suggesting that the parasite uses somewhat different machinery for cell proliferation. The study is important because it shows that kinesin-5 is not essential universally for cell division and suggests that there may be other non-canonical mechanisms and regulators. The general concept that kinesin-5 is essential for spindle pole formation and chromosome segregation during cell division is not true for Plasmodium, and this is of interest to a broad readership.

show abstract

“…The process requires ten or more rounds of DNA replication, segregation, and mitotic division to create a syncytial cell (sporoblast) with thousands of nuclei over a period of several days (Francia and Striepen, 2014;Gerald et al, 2011). The proper organization of nuclei into individual sporoblasts is organized by putative MTOCs (Roques et al, 2019;Sinden and Strong, 1978). As condensin has been shown to play an important role in organizing MTOCs (Kim et al, 2014) it may be that in the absence of condensin the endomitotic division is impaired and no sporozoites are formed.…”

Section: Discussionmentioning

confidence: 99%

Plasmodium condensin core subunits (SMC2/SMC4) mediate atypical mitosis and are essential for parasite proliferation and transmission

Pandey

Abel

Boucher

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

Condensin is a multi-subunit protein complex that regulates chromosome organization, segregation and condensation during cell division in eukaryotes. In Plasmodium spp., the causative agent of malaria, cell division is atypical and the role of condensin is unclear. Here we examine the role of SMC2 and SMC4, the core subunits of condensin during endomitosis in schizogony and endoreduplication in male gametogenesis. SMC2 and SMC4 localize at discrete foci during schizogony, and with a diffuse nuclear distribution during male gametogenesis. ChIP-seq analyses suggest a centromeric location of SMC2/SMC4 only during schizogony. Co-immunoprecipitation data reveal the presence of both condensin complex I and II during male gametogenesis, but only the SMC2/SMC4 heterodimer during schizogony. Finally, knockdown of smc2 and smc4 gene expression revealed their essential roles in parasite proliferation and transmission. This study shows that condensin core subunits (SMC2/SMC4) have differential complex and distinct functions at different stages of the parasite life cycle. RT, AAH and KGLR conceived and designed all experiments. RT, RP, SA, MB, RJW, MZ, ER, AF, DB, ED and SW performed the GFP tagging and conditional knockdown experiments; RRS performed liver stage imaging; RP, MZ, ED and RT performed protein pulldown experiments; ARB performed the mass spectroscopy; RP and DG performed the phylogenetic analysis and molecular dynamics; SA, RP XML, GB, TH, KGLR and RT performed the RNAseq and ChIP-Seq experiments; RP, SA, AAH, KGLR and RT analyzed the data; RP, SA, AAH, KGLR and RT wrote the manuscript and all others contributed to it.

show abstract

PlasmodiumcentrinPbCEN-4 localizes to the putative MTOC and is dispensable for malaria parasite proliferation

Cited by 33 publications

References 40 publications

Real-time dynamics of Plasmodium NDC80 reveals unusual modes of chromosome segregation during parasite proliferation

Real-time dynamics of Plasmodium NDC80 reveals unusual modes of chromosome segregation during parasite proliferation

Plasmodium bergheikinesin-5 associates with the spindle apparatus during cell division and is important for efficient production of infectious sporozoites

Plasmodium condensin core subunits (SMC2/SMC4) mediate atypical mitosis and are essential for parasite proliferation and transmission

Contact Info

Product

Resources

About