Nuclear Pore Complex Components in the Malaria Parasite Plasmodium berghei

Abstract

The nuclear pore complex (NPC) is a large macromolecular assembly of around 30 different proteins, so-called nucleoporins (Nups). Embedded in the nuclear envelope the NPC mediates bi-directional exchange between the cytoplasm and the nucleus and plays a role in transcriptional regulation that is poorly understood. NPCs display modular arrangements with an overall structure that is generally conserved among many eukaryotic phyla. However, Nups of yeast or human origin show little primary sequence conservation with those from early-branching protozoans leaving those of the malaria parasite unrecognized. Here we have combined bioinformatic and genetic methods to identify and spatially characterize Nup components in the rodent infecting parasite Plasmodium berghei and identified orthologs from the human malaria parasite P. falciparum, as well as the related apicomplexan parasite Toxoplasma gondii. For the first time we show the localization of selected Nups throughout the P. berghei life cycle. Largely restricted to apicomplexans we identify an extended C-terminal poly-proline extension in SEC13 that is essential for parasite survival and provide high-resolution images of Plasmodium NPCs obtained by cryo electron tomography. Our data provide the basis for full characterization of NPCs in malaria parasites, early branching unicellular eukaryotes with significant impact on human health.

CRISPR/Cas9-engineered inducible gametocyte producer lines as a novel tool for basic and applied research onPlasmodium falciparummalaria transmission stages

“…These observations are consistent with the open chromatin conformation and high transcriptional activity observed in both life cycle stages 52,90,91 and are suggestive of a high demand for nucleocytoplasmic shuttling during periods of growth. Importantly, because NPCs serve as a surrogate marker delineating the nuclear envelope our results also allow us to conclude that the nucleus in stage II to V gametocytes (i) undergoes marked morphological transformations reflected in narrow lateral extensions along the longitudinal axis of the gametocyte or rounded expansions of the nuclear envelope; and (ii) is frequently considerably larger than what might be extrapolated from the Hoechst-stained bulk of genetic material, in line with observations made in P. berghei gametocytes 81 . In some late stage gametocytes we even observed two distinct NPC-demarcated regions, of which one or both stained positive with Hoechst.…”

An extended DNA-free intranuclear compartment organizes centrosomal microtubules in Plasmodium falciparum

“…In absence of a known nuclear envelope marker for Plasmodium spp., we used a strain where the nuclear pore protein Nup313 has been tagged endogenously with 3xHA ( Fig. S3) to partly mark the nuclear boundary (Kehrer et al, 2018). Immunofluorescence co-staining with centrin and tubulin revealed that centrin signals localize on the cytoplasmic side, while microtubule ends are localized inside the nucleus during all stages of schizogony ( Fig.…”

CRISPR/Cas9-engineered inducible gametocyte producer lines as a novel tool for basic and applied research on Plasmodium falciparum malaria transmission stages

“…berghei gametocytes 81 . In some late stage gametocytes we even observed two distinct NPC-demarcated regions, of which one or both stained positive with Hoechst.…”

“…NPCs act as essential gateways for molecular transport into and out of the nucleus and play additional crucial roles in the regulation of gene expression and genome organization 77,78 . In malaria parasites, nuclear pores have hardly been studied and only six NUPs have so far been identi ed [79][80][81][82][83] , of which four have been localized in P. falciparum asexual blood stage parasites [PfNUP116 (PF3D7_1473700) 80 , PfNUP221/PfNUP100 (PF3D7_0905100) 83 , PfNUP313 (PF3D7_1446500) 79 , PfSEC13 (PF3D7_1230700) 82 ].…”