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Nucleopore complexes are essential in the maintenance of ploidy in Trypanosomatid parasites.
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International audience. Trypanosoma brucei and Leishmania spp. exhibit many original molecular and cellular features, among which a closed mitosis which remains poorly known. In terms of ploidy, while T. brucei is diploid, Leishmania spp. display mosaic aneuploidy aneuploidy demonstrated by FISH and confirmed by NGS. The biological significance of mosaic aneuploidy is still poorly understood but it has been associated with drug resistance and environment adaptability. Nuclear pore complexes (NPCs) are large multiprotein channels embedded in the nuclear envelope and made of nucleoporins. Although NPCs' primary function is to regulate bidirectional nucleocytoplasmic exchanges of macromolecules, recent research indicates that nucleoporins are involved in other core processes. Using tagged proteins, we determined that Mlp1 was located at the nuclear basket of the NPCs, while Mlp2 was intranuclear in interphasic cells and relocated at the poles of the mitotic spindle during mitosis. In T. brucei, RNAi knockdown of TbMlp2 had no effect on cellular growth whereas Mlp1 appears to be an essential gene. However, the use of flow cytometry and FISH revealed that both RNAi knockdowns perturbed the distribution of chromosomes during mitosis, leading to aneuploidy. The phenotypes observed here are in agreement with the hypothesis that Mlp2 plays a role in chromosomal segregation and might be involved in the constitution of kinetochores while Mlp1 would play a role in the regulation of replication. Indeed, in TbMlp1 knockdown, the vast majority of cells divided asymmetrically and in all dividing nuclei, the total number of homologs was an odd number (i.e: 1:2 or 2:3).The role of Mlp1 in DNA replication will be further explored by DNA molecular combing and Short Nascent Strand Sequencing (SNS-seq).
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