Sialidases remove sialic acid residues from various sialo-derivatives. To gain further insights into the biological roles of sialidases in vertebrates, we exploited zebrafish (Danio rerio) as an animal model. A zebrafish transcriptome- and genome-wide search using the sequences of the human NEU polypeptides as templates revealed the presence of seven different genes related to human sialidases. neu1 and neu4 are the putative orthologues of the mammalian sialidases NEU1 and NEU4 respectively. Interestingly, the remaining genes are organized in clusters located on chromosome 21 and are all more closely related to mammalian sialidase NEU3. They were thus named neu3.1, neu3.2, neu3.3, neu3.4 and neu3.5. Using RT–PCR (reverse transcription–PCR) we detected transcripts for all genes, apart from neu3.4, and whole-mount in situ hybridization experiments show a localized expression pattern in gut and lens for neu3.1 and neu4 respectively. Transfection experiments in COS7 (monkey kidney) cells demonstrate that Neu3.1, Neu3.2, Neu3.3 and Neu4 zebrafish proteins are sialidase enzymes. Neu3.1, Neu3.3 and Neu4 are membrane-associated and show a very acidic pH optimum below 3.0, whereas Neu3.2 is a soluble sialidase with a pH optimum of 5.6. These results were further confirmed by subcellular localization studies carried out using immunofluorescence. Moreover, expression in COS7 cells of these novel zebrafish sialidases (with the exception of Neu3.2) induces a significant modification of the ganglioside pattern, consistent with the results obtained with membrane-associated mammalian sialidases. Overall, the redundancy of sialidases together with their expression profile and their activity exerted on gangliosides of living cells indicate the biological relevance of this class of enzymes in zebrafish.
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December 2007
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Research Article|
November 28 2007
Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio)
Marta Manzoni;
Marta Manzoni
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
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Paolo Colombi;
Paolo Colombi
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
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Nadia Papini;
Nadia Papini
†Department of Medical Chemistry, Biochemistry and Biotechnology, L.I.T.A. (Laboratorio Interdisciplinare di Tecnologie Avanzate)-Segrate, School of Medicine, University of Milano, via Fratelli Cervi 93, 20090 Segrate, Italy
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Luana Rubaga;
Luana Rubaga
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
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Natascia Tiso;
Natascia Tiso
‡Department of Biology, University of Padova, via U. Bassi 58/B, 35131 Padova, Italy
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Augusto Preti;
Augusto Preti
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
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Bruno Venerando;
Bruno Venerando
†Department of Medical Chemistry, Biochemistry and Biotechnology, L.I.T.A. (Laboratorio Interdisciplinare di Tecnologie Avanzate)-Segrate, School of Medicine, University of Milano, via Fratelli Cervi 93, 20090 Segrate, Italy
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Guido Tettamanti;
Guido Tettamanti
†Department of Medical Chemistry, Biochemistry and Biotechnology, L.I.T.A. (Laboratorio Interdisciplinare di Tecnologie Avanzate)-Segrate, School of Medicine, University of Milano, via Fratelli Cervi 93, 20090 Segrate, Italy
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Roberto Bresciani;
Roberto Bresciani
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
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Francesco Argenton;
Francesco Argenton
‡Department of Biology, University of Padova, via U. Bassi 58/B, 35131 Padova, Italy
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Giuseppe Borsani;
Giuseppe Borsani
1
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
1Correspondence may be addressed to either of these authors (email gborsani@med.unibs.it or monti@med.unibs.it)
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Eugenio Monti
Eugenio Monti
1
*Department of Biomedical Sciences and Biotechnology, School of Medicine, University of Brescia, viale Europa 11, 25123 Brescia, Italy
1Correspondence may be addressed to either of these authors (email gborsani@med.unibs.it or monti@med.unibs.it)
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Publisher: Portland Press Ltd
Received:
May 10 2007
Revision Received:
August 02 2007
Accepted:
August 21 2007
Accepted Manuscript online:
August 21 2007
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2007 Biochemical Society
2007
Biochem J (2007) 408 (3): 395–406.
Article history
Received:
May 10 2007
Revision Received:
August 02 2007
Accepted:
August 21 2007
Accepted Manuscript online:
August 21 2007
Citation
Marta Manzoni, Paolo Colombi, Nadia Papini, Luana Rubaga, Natascia Tiso, Augusto Preti, Bruno Venerando, Guido Tettamanti, Roberto Bresciani, Francesco Argenton, Giuseppe Borsani, Eugenio Monti; Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio). Biochem J 15 December 2007; 408 (3): 395–406. doi: https://doi.org/10.1042/BJ20070627
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