Glycobiology Advance Access published online on June 19, 2007
Glycobiology, doi:10.1093/glycob/cwm064
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The rainbow trout CMP-sialic acid synthetase utilises a nuclear localization signal different from that identified in the mouse enzyme
1 Institute for Glycomics, Griffith University (Gold Coast Campus), PMB 50 Gold Coast Mail Centre, 9726, QLD Australia
2 Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan
3 Abteilung Zelluläre Chemie, Zentrum Biochemie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, D-30625 Hannover, Germany
4 To whom correspondence should be addressed: Dr Anja K. Münster-Kühnel, Abteilung Zelluläre Chemie, Zentrum Biochemie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, D-30625 Hannover, Germany. Phone: ++49 511 532 3367; Fax: ++49 511 532 3956; email: Muenster.Anja{at}mh-hannover.de
Received on April 19, 2007; accepted on June 7, 2007
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 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 localisation 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 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 localisation of the enzyme, provides supporting evidence that nuclear localisation is linked to a second yet unknown function.
Key words: CMP-sialic acid / CMP-sialic acid synthetase / nuclear localization signal / sialic acids