Transcriptional regulation of the human GD3 synthase gene expression in Fas-induced Jurkat T cells: a critical role of transcription factor NF-kappaB in regulated expression.
The transcriptional regulation mechanisms involved in the up-regulation of Fas-induced GD3 synthase gene have not yet been elucidated. 5'-Rapid amplification of cDNA end (5'-RACE) using mRNA prepared from Fas-induced Jurkat T cells revealed the presence of multiple transcription start sites of human GD3 synthase gene, and the 5'-end analysis of the longest of its product showed that transcription started from 650 nucleotides upstream of the translational initiation site. Promoter analyses of the 5'-flanking region of the human GD3 synthase gene using luciferase gene reporter system showed strong promoter activity in Fas-induced Jurkat T cells. Deletion study revealed that the region from -1146 to -646 (A of the translational start ATG as position +1) was indispensable for the Fas response. This region lacks apparent TATA and CAAT boxes but contains putative binding sites for transcription factors c-Ets-1, cAMP-responsive element-binding (CREB) protein, activating protein 1 (AP-1), and NF-kappaB. Base-substitution experiment showed that only the NF-kappaB-binding site of putative binding sites is required for the maximal expression induced by Fas. Both DNase I footprint and electrophoretic mobility shift assays with the nuclear extract of Fas-induced Jurkat T cells revealed that NF-kappaB was bound specifically to the probe being mediated by its binding site in the promoter sequence. Taken together, these results indicate that NF-kappaB plays an essential role in the transcriptional activity of human GD3 synthase gene in Fas-induced Jurkat T cells. In addition, the translocation of NF-kappaB-binding protein to nucleus by Fas activation is also crucial for the increased expression of the GD3 synthase gene in Fas-activated Jurkat T cells.
Systematic Proteomic Research Center, Korea Research Institute of Bioscience and Biotechnology, Yusong-Gu, Daejon 305-600, Korea.
Nam-Young
NY
Kang
Young
Y
Kang
Sung-Koo
SK
Kang
Young-Choon
YC
Lee
Hee-Jeong
HJ
Choi
Young-Seek
YS
Lee
Soo-Young
SY
Cho
Yong-Sam
YS
Kim
Jeong-Heon
JH
Ko
Cheorl-Ho
CH
Kim
D001483:Base Sequence; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D012799:Sialyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013601:T-Lymphocytes / Q000187:drug effects* / Q000201:enzymology*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000494:pharmacology*
D016328:NF-kappa B; D012333:RNA, Messenger; D019014:fas Receptor; D012799:Sialyltransferases; C043662:alpha-N-acetylneuraminate alpha-2,8-sialyltransferase