title: NLS copy-number variation governs efficiency of nuclear import--case study on dUTPases.
abstract: Nucleocytoplasmic trafficking of large macromolecules requires an active transport machinery. In many cases, this is initiated by binding of the nuclear localization signal (NLS) peptide of cargo proteins to importin-α molecules. Fine orchestration of nucleocytoplasmic trafficking is of particularly high importance for proteins involved in maintenance of genome integrity, such as dUTPases, which are responsible for prevention of uracil incorporation into the genome. In most eukaryotes, dUTPases have two homotrimeric isoforms: one of these contains three NLSs and is present in the cell nucleus, while the other is located in the cytoplasm or the mitochondria. Here we focus on the unusual occurrence of a pseudo-heterotrimeric dUTPase in Drosophila virilis that contains one NLS, and investigate its localization pattern compared to the homotrimeric dUTPase isoforms of Drosophila melanogaster. Although the interaction of individual NLSs with importin-α has been well characterized, the question of how multiple NLSs of oligomeric cargo proteins affect their trafficking has been less frequently addressed in adequate detail. Using the D. virilis dUTPase as a fully relevant physiologically occurring model protein, we show that NLS copy number influences the efficiency of nuclear import in both insect and mammalian cell lines, as well as in D. melanogaster and D. virilis tissues. Biophysical data indicate that NLS copy number determines the stoichiometry of complexation between importin-α and dUTPases. The main conclusion of our study is that, in D. virilis, a single dUTPase isoform efficiently reproduces the cellular dUTPase distribution pattern that requires two isoforms in D. melanogaster.
authors.affiliation: Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary; Department of Applied Biotechnology and Food Sciences, Budapest University of Technology and Economics, Hungary.
authors.forename: Gergely
authors.initials: G
authors.lastname: Róna
authors.forename: Hajnalka L
authors.initials: HL
authors.lastname: Pálinkás
authors.forename: Máté
authors.initials: M
authors.lastname: Borsos
authors.forename: András
authors.initials: A
authors.lastname: Horváth
authors.forename: Ildikó
authors.initials: I
authors.lastname: Scheer
authors.forename: András
authors.initials: A
authors.lastname: Benedek
authors.forename: Gergely N
authors.initials: GN
authors.lastname: Nagy
authors.forename: Imre
authors.initials: I
authors.lastname: Zagyva
authors.forename: Beáta G
authors.initials: BG
authors.lastname: Vértessy
mesh_terms: D000818:Animals; D001704:Biopolymers / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism*; D056915:DNA Copy Number Variations; D004330:Drosophila; D019913:Nuclear Localization Signals / Q000235:genetics*; D021381:Protein Transport
chemical_list: D001704:Biopolymers; D019913:Nuclear Localization Signals
keywords: dUTPase; importin; nuclear localization signal; nucleocytoplasmic trafficking; oligomer proteins; regulation; stoichiometry