title: The rainbow trout CMP-sialic acid synthetase utilises a nuclear localization signal different from that identified in the mouse enzyme.
abstract: The terminal sugar sialic acid (Sia) plays a pivotal role in cell-cell interaction and recognition. A prerequisite for the biosynthesis of sialoglycoconjugates is the activation of Sia to cytidine monophosphate-Sia (CMP-Sia), by CMP-Sia synthetases (CMP-Sia-syn). CMP-Sia-syn are conserved from bacteria to man, and have been found to reside in the nucleus of all vertebrate species analysed to date. We previously cloned the CMP-Sia-syn from rainbow trout (rt) and identified three clusters of basic amino acids (BC) that might act as nuclear localization signals (NLS). Here, we utilised chimeric proteins and rt CMP-Sia-syn mutants in which putative NLS sequences were deleted, to identify the nuclear transport signal. Divergent from the mouse enzyme, where the crucial NLS is part of the enzyme's active site, in the rt CMP-Sia-syn the NLS and active site are disparate. The crucial NLS in the fish enzyme is bipartite and the functionality depends on a free N-terminus. Comparative analysis of all putative rt NLS in mouse and fish cells identified a second inferior motif (rtBC5-6), which was functional only in fish cells suggesting some differences in transport mechanism or folding variabilities in fish. Moreover, based on computational analyses of putative CMP-Sia-syn from distant deuterostomian organisms it was concluded that CMP-Sia-syn nuclear localization is a relatively recent invention, originating in echinoderms. In summary, our data describing structural differences in the NLS of vertebrate CMP-Sia-syn, and the independence of Sia activation from the subcellular localization of the enzyme, provide supporting evidence that nuclear localization is linked to a second yet unknown function.
authors.affiliation: Institute for Glycomics, Griffith University (Gold Coast Campus), PMB 50 Gold Coast Mail Centre 9726, Gold Coast, QLD, Australia.
authors.forename: Joe
authors.initials: J
authors.lastname: Tiralongo
authors.forename: Akiko
authors.initials: A
authors.lastname: Fujita
authors.forename: Chihiro
authors.initials: C
authors.lastname: Sato
authors.forename: Ken
authors.initials: K
authors.lastname: Kitajima
authors.forename: Friederike
authors.initials: F
authors.lastname: Lehmann
authors.forename: Melanie
authors.initials: M
authors.lastname: Oschlies
authors.forename: Rita
authors.initials: R
authors.lastname: Gerardy-Schahn
authors.forename: Anja K
authors.initials: AK
authors.lastname: Münster-Kühnel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid / Q000737:chemistry; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D000216:N-Acylneuraminate Cytidylyltransferase / Q000737:chemistry* / Q000378:metabolism*; D041681:NIH 3T3 Cells; D019913:Nuclear Localization Signals; D017686:Oncorhynchus mykiss; D015398:Signal Transduction; D013045:Species Specificity
chemical_list: D019913:Nuclear Localization Signals; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid; D000216:N-Acylneuraminate Cytidylyltransferase