Valproic acid-mediated transcriptional regulation of human GM3 synthase (hST3Gal V) in SK-N-BE(2)-C human neuroblastoma cells.
OBJECTIVE
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 immunofluorescent 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, sitedirected mutagenesis and electrophoretic mobility shift assay indicated that the CRE at -143 is crucial for the VPA-induced expression of hST3Gal V in SK-NBE( 2)-C cells.


CONCLUSIONS
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.
Kwon
Haw-young
HY
Department of Biotechnology, Dong-A University, Busan 604-714, Republic of Korea.
Kang
Nam-young
NY
Dae
Hyun-mi
HM
Kim
Kyoung-sook
KS
Kim
Cheorl-ho
CH
Do
Su-il
SI
Lee
Youngchoon
Y
D000927:Anticonvulsants / Q000494:pharmacology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival; D005679:G(M3) Ganglioside / Q000096:biosynthesis; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D008156:Luciferases / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012799:Sialyltransferases / Q000096:biosynthesis*; D014162:Transfection; D014635:Valproic Acid / Q000494:pharmacology*
D000927:Anticonvulsants; D005679:G(M3) Ganglioside; D012333:RNA, Messenger; D014635:Valproic Acid; D008156:Luciferases; D012799:Sialyltransferases; C021343:haematoside synthetase