Nucleoside diphosphate kinase beta (Nm23-R1/NDPKbeta) is associated with intermediate filaments and becomes upregulated upon cAMP-induced differentiation of rat C6 glioma.
Nucleoside diphosphate kinases (Nm23/NDPK) are enzymes functional in cell proliferation, differentiation, development, tumor progression, and metastasis. Nevertheless, no consensus exists about the molecular mechanism by which Nm23/NDPK isoforms exert their role in these processes. We investigated the expression of the rat Nm23-R1/NDPKbeta and Nm23-R2/NDPKalpha isoforms, homologues of the human Nm23-H1/NDPK ... A and Nm23-H2/NDPK B proteins, respectively, upon cAMP-induced differentiation of rat C6 glioma cells and demonstrated a differential interaction with intermediate filaments. Semiquantitative RT-PCR, immunoblotting, and flow cytometry showed a constitutive expression of both Nm23 isoforms. After induction of differentiation in C6 cells with cAMP analogs or isoproterenol, a dose-dependent 2- and 2.5-fold upregulation of the Nm23-R1 mRNA and protein, respectively, was observed. In contrast, the expression of Nm23-R2 remained unchanged. Localization of both isoforms with confocal laser scanning microscopy demonstrated a punctate reticular staining pattern for both Nm23 isoforms in the cytosol and processes of the cells which was particularly intense in the perinuclear region. In addition, while Nm23-R2 was colocalized and coimmunoprecipitated with vimentin in nondifferentiated cells, both isoforms were associated with GFAP in differentiated cells. The significance of these findings in relation to a possible function of Nm23 isoforms in cell proliferation, differentiation, and tumor-associated mechanisms is discussed.
Mesh Terms:
8-Bromo Cyclic Adenosine Monophosphate, Animals, Bucladesine, Cell Differentiation, Cyclic AMP, Cytosol, Gene Expression Regulation, Enzymologic, Glioma, Humans, Intermediate Filaments, Isoenzymes, Isoproterenol, Kinetics, Monomeric GTP-Binding Proteins, NM23 Nucleoside Diphosphate Kinases, Nucleoside-Diphosphate Kinase, Protein Biosynthesis, Rats, Reverse Transcriptase Polymerase Chain Reaction, Transcription Factors, Transcription, Genetic, Tumor Cells, Cultured
8-Bromo Cyclic Adenosine Monophosphate, Animals, Bucladesine, Cell Differentiation, Cyclic AMP, Cytosol, Gene Expression Regulation, Enzymologic, Glioma, Humans, Intermediate Filaments, Isoenzymes, Isoproterenol, Kinetics, Monomeric GTP-Binding Proteins, NM23 Nucleoside Diphosphate Kinases, Nucleoside-Diphosphate Kinase, Protein Biosynthesis, Rats, Reverse Transcriptase Polymerase Chain Reaction, Transcription Factors, Transcription, Genetic, Tumor Cells, Cultured
Exp. Cell Res.
Date: Nov. 25, 2000
PubMed ID: 11082283
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