title: Characterization of a nuclear pore protein sheds light on the roles and composition of the Toxoplasma gondii nuclear pore complex.
abstract: The nuclear pore is a key structure in eukaryotes regulating nuclear-cytoplasmic transport as well as a wide range of cellular processes. Here, we report the characterization of the first Toxoplasma gondii nuclear pore protein, named TgNup302, which appears to be the orthologue of the mammalian Nup98-96 protein. We produced a conditional knock-down mutant that expresses TgNup302 under the control of an inducible tetracycline-regulated promoter. Under ATc treatment, a substantial decrease of TgNup302 protein in inducible knock-down (iKD) parasites was observed, causing a delay in parasite proliferation. Moreover, the nuclear protein TgENO2 was trapped in the cytoplasm of ATc-treated mutants, suggesting that TgNup302 is involved in nuclear transport. Fluorescence in situ hybridization revealed that TgNup302 is essential for 18S RNA export from the nucleus to the cytoplasm, while global mRNA export remains unchanged. Using an affinity tag purification combined with mass spectrometry, we identified additional components of the nuclear pore complex, including proteins potentially interacting with chromatin. Furthermore, reverse immunoprecipitation confirmed their interaction with TgNup302, and structured illuminated microscopy confirmed the NPC localization of some of the TgNup302-interacting proteins. Intriguingly, facilitates chromatin transcription complex (FACT) components were identified, suggesting the existence of an NPC-chromatin interaction in T. gondii. Identification of TgNup302-interacting proteins also provides the first glimpse at the NPC structure in Apicomplexa, suggesting a structural conservation of the NPC components between distant eukaryotes.
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France.
authors.forename: Flavie
authors.initials: F
authors.lastname: Courjol
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France.
authors.forename: Thomas
authors.initials: T
authors.lastname: Mouveaux
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France.
authors.forename: Kevin
authors.initials: K
authors.lastname: Lesage
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France.
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Saliou
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Werkmeister
authors.affiliation: MGX-Montpellier GenomiX, c/o Institut de Génomique Fonctionnelle, 141 rue de la cardonille, 34094, Montpellier Cedex 5, France.
authors.forename: Maurine
authors.initials: M
authors.lastname: Bonabaud
authors.affiliation: MGX-Montpellier GenomiX, c/o Institut de Génomique Fonctionnelle, 141 rue de la cardonille, 34094, Montpellier Cedex 5, France.
authors.forename: Marine
authors.initials: M
authors.lastname: Rohmer
authors.affiliation: Laboratory of Cell Physiology, INSERM U 1003, Université Lille Nord de France, Villeneuve d'Ascq, France.
authors.forename: Christian
authors.initials: C
authors.lastname: Slomianny
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France.
authors.forename: Franck
authors.initials: F
authors.lastname: Lafont
authors.affiliation: University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, CIIL-Centre d'Infection et d'Immunité de Lille, 59000, Lille, France. mathieu.gissot@pasteur-lille.fr.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Gissot
mesh_terms: D000818:Animals; D064113:CRISPR-Cas Systems; D002467:Cell Nucleus / Q000378:metabolism; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D059014:High-Throughput Nucleotide Sequencing; D008853:Microscopy; D022022:Nuclear Pore / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins / Q000378:metabolism*; D010271:Parasites / Q000378:metabolism; D010641:Phenotype; D011485:Protein Binding; D021381:Protein Transport; D015800:Protozoan Proteins / Q000378:metabolism*; D034443:RNA Transport; D012337:RNA, Ribosomal, 18S / Q000378:metabolism; D014122:Toxoplasma / Q000254:growth & development / Q000378:metabolism*
chemical_list: D028861:Nuclear Pore Complex Proteins; D015800:Protozoan Proteins; D012337:RNA, Ribosomal, 18S
keywords: Apicomplexa; Gene expression; Nuclear pore complex; Nucleoporins; Toxoplasma gondii