Manzoni, M., Colombi, P., Papini, N., Rubaga, L., Tiso, N., Preti, A., Venerando, B., Tettamanti, G., Bresciani, R., Argenton, F., Borsani, G., and Monti, E. (2007) Molecular cloning and biochemical characterization of sialidases from zebrafish (Danio rerio). The Biochemical journal. 408(3):395-406.
Sialidases remove sialic acid residues from various sialo-derivatives. To gain further insights into the biological roles of sialidases in vertebrates, we exploited zebrafish as animal model. A zebrafish transcriptome- and genome-wide search using as a query the sequences of the human NEU polypeptides reveal the presence of 7 different genes related to human sialidases. While neu1 and neu4 are the putative orthologs of the mammalian sialidases NEU1 and NEU4, respectively, the remaining genes are organized in a clusters located on chromosome 21 and they are all more closely related to mammalian sialidase NEU3. They were thus named neu3.1, neu3.2, neu3.3, neu3.4 and neu3.5. By RT-PCR we detect transcripts for all genes but 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 cells demonstrate that Neu3.1, Neu3.2, Neu3.3 and Neu4 polypeptides 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 data are further confirmed by subcellular localization studies carried out by immunofluorescence. Moreover, expression of these novel zebrafish sialidases but Neu3.2 induces a significant modification of the ganglioside pattern of living COS7 cells, consistent with the results obtained in the case of membrane-associated mammalian enzymes. 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.