Original Article

Valproic acid-mediated transcriptional regulation of human GM3 synthase (hST3Gal V) in SK-N-BE(2)-C human neuroblastoma cells

Authors: Haw-young Kwon, Nam-young Kang, Hyun-mi Dae, Kyoung-sook Kim, Cheorl-ho Kim, Su-il Do, Young-choon Lee

Abstract

Aim: To investigate whether valproic acid (VPA) modulates human GM3 synthase (hST3Gal V) mRNA expression, as a part of ganglioside GM3 biosynthesis, in human neuroblastoma cells.
Methods: Using RT-PCR and immunofluo-rescent confocal microscopy, we examined hST3Gal V mRNA and GM3 levels during VPA-induced differentiation of human neuroblastoma SK-N-BE(2)-C cells. We characterized the VPA-inducible promoter region within the hST3-Gal V gene using luciferase constructs carrying 5′-deletions of the hST3Gal V promoter.
Results: RT-PCR indicated that VPA-mediated hST3Gal V induction is transcriptionally regulated. Functional analysis of the 5′-flanking region of the hST3Gal V gene demonstrated that the -177 to -83 region, which contains a cAMP-responsive element (CRE) at -143, functions as the VPA-inducible promoter by actively binding CRE binding protein (CREB). In addition, site-directed mutagenesis and electrophoretic mobility shift assay indicated that the CRE at -143 is crucial for the VPA-induced expression of hST3Gal V in SK-N-BE(2)-C cells.
Conclusion: Our results isolated the core promoter region in the hST3Gal V promoter, a CRE at -143, and demonstrated that it is essential for transcriptional activation of hST3Gal V in VPA-induced SK-N-BE(2)-C cells. Subsequent CREB binding to this CRE mediates VPA-dependent upregulation of hST3Gal V gene expression.
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