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NLS copy‐number variation governs efficiency of nuclear import – case study on d UTP ases
Author(s) -
Róna Gergely,
Pálinkás Hajnalka L.,
Borsos Máté,
Horváth András,
Scheer Ildikó,
Benedek András,
Nagy Gergely N.,
Zagyva Imre,
Vértessy Beáta G.
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.13086
Subject(s) - nuclear transport , nuclear localization sequence , nls , importin , microbiology and biotechnology , nucleoplasm , cytoplasm , biology , gene isoform , transport protein , cell nucleus , genetics , gene , nucleolus
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 d UTP ases, which are responsible for prevention of uracil incorporation into the genome. In most eukaryotes, d UTP ases have two homotrimeric isoforms: one of these contains three NLS s 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 d UTP ase in Drosophila virilis that contains one NLS , and investigate its localization pattern compared to the homotrimeric d UTP ase isoforms of Drosophila melanogaster . Although the interaction of individual NLS s with importin‐α has been well characterized, the question of how multiple NLS s of oligomeric cargo proteins affect their trafficking has been less frequently addressed in adequate detail. Using the D. virilis d UTP ase 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 d UTP ases. The main conclusion of our study is that, in D. virilis , a single d UTP ase isoform efficiently reproduces the cellular d UTP ase distribution pattern that requires two isoforms in D. melanogaster . Structured digital abstract • ABC dUTPase and importin-a bind by molecular sieving ( View interaction ) • AAA dUTPase and importin-a bind by molecular sieving ( View interaction ) • ABC dUTPase and importin-a bind by comigration in non denaturing gel electrophoresis ( View interaction ) • AAA dUTPase and importin-a bind by comigration in non denaturing gel electrophoresis ( View interaction ) • ABC dUTPase and importin-a bind by isothermal titration calorimetry ( View interaction ) • AAA dUTPase and importin-a bind by isothermal titration calorimetry ( View interaction )