Molecular cloning of mouse ganglioside sialidase and its increased expression in Neuro2a cell differentiation

Takafumi Hasegawa, Kazunori Yamaguchi, Tadashi Wada, Atsushi Takeda, Yasuto Itoyama, Taeko Miyagi

Research output: Contribution to journalArticle

113 Citations (Scopus)

Abstract

Ganglioside sialidases have been implicated in neuronal differentiation processes, including neurite outgrowth. To understand further the roles and regulation mechanisms of the sialidase in neuronal systems, we have cloned mouse ganglioside sialidase cDNA and observed its expression in Neuro2a cell differentiation. A 3339-base pair cDNA, cloned based on the sequence information of previously cloned enzymes, encodes 418 amino acids containing three Asp boxes characteristic of sialidases. Northern blot analysis revealed a 3.4-kilobase transcript expressed highly in heart but also in several other tissues including brain. In situ hybridization of mouse brain demonstrated the mRNA to be present in the cerebral cortex, as well as in the granule cell layer, Purkinje cells, and deep cerebellar nucleus of the cerebellum. Transient expression of the cDNA in COS-1 cells resulted in over 300-fold increase in sialidase activity toward gangliosides compared with the control level, with a preference for ganglioside substrate. During 5- bromodeoxyuridine-induced Neuro2a cell differentiation, the expression of the sialidase was increased as assessed by activity assays and quantitative reverse transcription-polymerase chain reaction analyses. Stable transfection of the sialidase in Neuro2a cells resulted in accelerated neurite arborization following 5-bromodeoxyuridine treatment, indicating the direct participation of this ganglioside sialidase in neuronal cell differentiation.

Original languageEnglish
Pages (from-to)8007-8015
Number of pages9
JournalJournal of Biological Chemistry
Volume275
Issue number11
DOIs
Publication statusPublished - 2000 Mar 17

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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