Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio)

Marta Manzoni, Paolo Colombi, Nadia Papini, Luana Rubaga, Natascia Tiso, Augusto Preti, Bruno Venerando, Guido Tettamanti, Roberto Bresciani, Francesco Argenton, Giuseppe Borsani, Eugenio Monti

Research output: Contribution to journalArticle

Abstract

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.

Original languageEnglish
Pages (from-to)395-406
Number of pages12
JournalBiochemical Journal
Volume408
Issue number3
DOIs
Publication statusPublished - Dec 15 2007

Fingerprint

Cloning
Molecular Cloning
Zebrafish
Neuraminidase
Genes
Gangliosides
Zebrafish Proteins
Membranes
N-Acetylneuraminic Acid
Enzymes
Transcription
Chromosomes
Chromosomes, Human, Pair 21
Redundancy
Lenses
Animals
Transcriptome
Experiments
Cells
Reverse Transcription

Keywords

  • Comparative genomics
  • Evolution
  • Neuraminidase
  • Sialidase
  • Zebrafish

ASJC Scopus subject areas

  • Biochemistry

Cite this

Manzoni, M., Colombi, P., Papini, N., Rubaga, L., Tiso, N., Preti, A., ... Monti, E. (2007). Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio). Biochemical Journal, 408(3), 395-406. https://doi.org/10.1042/BJ20070627

Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio). / Manzoni, Marta; Colombi, Paolo; Papini, Nadia; Rubaga, Luana; Tiso, Natascia; Preti, Augusto; Venerando, Bruno; Tettamanti, Guido; Bresciani, Roberto; Argenton, Francesco; Borsani, Giuseppe; Monti, Eugenio.

In: Biochemical Journal, Vol. 408, No. 3, 15.12.2007, p. 395-406.

Research output: Contribution to journalArticle

Manzoni, M, Colombi, P, Papini, N, Rubaga, L, Tiso, N, Preti, A, Venerando, B, Tettamanti, G, Bresciani, R, Argenton, F, Borsani, G & Monti, E 2007, 'Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio)', Biochemical Journal, vol. 408, no. 3, pp. 395-406. https://doi.org/10.1042/BJ20070627
Manzoni, Marta ; Colombi, Paolo ; Papini, Nadia ; Rubaga, Luana ; Tiso, Natascia ; Preti, Augusto ; Venerando, Bruno ; Tettamanti, Guido ; Bresciani, Roberto ; Argenton, Francesco ; Borsani, Giuseppe ; Monti, Eugenio. / Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio). In: Biochemical Journal. 2007 ; Vol. 408, No. 3. pp. 395-406.
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