Research Article: Identification of Plasmodium falciparum nuclear proteins by mass spectrometry and proposed protein annotation

Date Published: October 31, 2018

Publisher: Public Library of Science

Author(s): Sylvie Briquet, Asma Ourimi, Cédric Pionneau, Juliana Bernardes, Alessandra Carbone, Solenne Chardonnet, Catherine Vaquero, Gordon Langsley.

http://doi.org/10.1371/journal.pone.0205596

Abstract

The nuclear proteome of Plasmodium falciparum results from the continual shuttle of proteins between the cell cytoplasm—nucleus and vice versa. Using shotgun proteomics tools, we explored the nuclear proteins of mixed populations of Plasmodium falciparum extracted from infected erythrocytes. We combined GeLC-MS/MS and 2D-LC-MS/MS with a peptide ion exclusion procedure in order to increase the detection of low abundant proteins such as those involved in gene expression. We have identified 446 nuclear proteins covering all expected nuclear protein families involved in gene regulation. All structural ribosomal (40S and 60S) proteins were identified which is consistent with the nuclear localization of ribosomal biogenesis. Proteins involved in the translation machinery were also found suggesting that translational events might occur in the nucleus in P. falciparum as previously hypothesized in eukaryotes. These data were compared to the protein list established by PlasmoDB and submitted to Plasmobase a recently reported Plasmodium annotation website to propose new functional putative annotation of several unknown proteins found in the nuclear extracts.

Partial Text

In eukaryote cells, the nucleus is a highly dynamic and complex organelle [1] [2] where major regulatory gene expression events take place such as DNA replication, RNA synthesis within transcriptional machinery, mRNA processing and transport to the cytoplasm as well as ribosomal sub-units biogenesis. The nucleus is also organized to participate in RNA, protein and ribosomal sub-unit trafficking in and out of the nucleus [3]. In Plasmodium, the dynamic organization and function of the nucleus vary throughout the different stages of cell development from active multiplication to parasite differentiation. Diverse sub-proteomes of the nucleus such as nucleolus or nuclear membranes have been investigated in eukaryotes by a number of proteomics experimental procedures including 2D gels, 1D gels followed by LC-MS/MS (GeLC-MS/MS), direct 2D-LC-MS/MS shotgun analysis or a combination of these approaches. A combination of both experimental approaches enhances the coverage compared to either individual methods [4]. However, a comprehensive identification of nuclear proteins still awaits completion in eukaryotes. Of note, a continuous cytoplasm-nucleus protein shuttling occurs in eukaryotes weakening the characterisation and definition of nuclear proteins even though proteins accumulate either in the cytoplasm or in the nucleus according to the cellular development [5]. This back and forth protein transport through the nuclear pore plays an important role in the control of gene expression.

Our aim was to increase our knowledge about the protein content of the nucleus in Plasmodium falciparum. In this attempt, we conducted a large-scale proteomic analysis of 3D7 P. falciparum nuclear extracts obtained from unsynchronized cultures of parasitized erythrocytic cells. The use of mixed populations of the parasite gives access to all nuclear proteins whatever the stage.

 

Source:

http://doi.org/10.1371/journal.pone.0205596

 

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