An extended DNA-free intranuclear compartment organizes centrosome microtubules in malaria parasites

Abstract

Proliferation of Plasmodium falciparum in red blood cells is the cause of malaria and is underpinned by an unconventional cell division mode, called schizogony. Contrary to model organisms, P. falciparum replicates by multiple rounds of nuclear divisions that are not interrupted by cytokinesis. Organization and dynamics of critical nuclear division factors remain poorly understood. Centriolar plaques, the centrosomes of P. falciparum, serve as microtubule organizing centers and have an acentriolar, amorphous structure. The small size of parasite nuclei has precluded detailed analysis of intranuclear microtubule organization by classical fluorescence microscopy. We apply recently developed super-resolution and time-lapse imaging protocols to describe microtubule reconfiguration during schizogony. Analysis of centrin, nuclear pore, and microtubule positioning reveals two distinct compartments of the centriolar plaque. Whereas centrin is extranuclear, we confirm by correlative light and electron tomography that microtubules are nucleated in a previously unknown and extended intranuclear compartment, which is devoid of chromatin but protein-dense. This study generates a working model for an unconventional centrosome and enables a better understanding about the diversity of eukaryotic cell division.

Expansion microsopy reveals Plasmodium falciparum blood-stage parasites undergo anaphase with a chromatin bridge in the absence of mini-chromosome maintenance complex binding protein

“…Ultrastructure Expansion Microscopy (U-ExM) was performed as previously described 149 [16,22,35], with significant modification. 150 12 mm round coverslips (Fisher Cat.…”

“…Despite the importance of nuclear envelope dynamics during 360 schizogony, and Plasmodium mitosis, there is currently no reliable marker of the P. falciparum 361 nuclear envelope for microscopic visualization. Previous studies have localized a few nucleoporins (nups) to the nuclear envelope of either P. falciparum [16,38] or P. berghei [39], but their distribution and number is dynamic across the lifecycle, limiting the robustness of nups as nuclear envelope markers. Therefore, we wanted to identify a uniform, U-ExM compatible stain for the P. falciparum nuclear envelope to allow us to study nuclear envelope changes in the context of MCMBP deficient parasites.…”

“…MTOCs [16]. The nomenclature around this microtubule structure is inconsistent, but in this 57 study, they will be referred to as interpolar spindles.…”

“…It has been shown that around when DNA replication begins, 51 the hemispindle retracts, the MTOC duplicates, and the mitotic spindle is formed. Mitotic 52 spindles consist of short microtubules that extend from two opposing MTOCs and connect to 53 the kinetochore, in preparation for chromatid separation into daughter nuclei [15,16,[18][19][20][21].…”

“…The nomenclature around this microtubule structure is inconsistent, but in this 57 study, they will be referred to as interpolar spindles. Following nuclear segregation, the 58 interpolar spindle retracts and the two daughter nuclei undergo nuclei fission before reforming 59 the hemispindle [16]. 60 Mitosis in Plasmodium has been poorly studied historically.…”