VIP gene transcription is regulated by far upstream enhancer and repressor elements.
SK-N-SH human neuroblastoma subclones differ widely in basal and second messenger induction of the gene encoding the neuropeptide vasoactive intestinal peptide (VIP). These differences were recapitulated by a chimeric gene which consisted of 5.2 kb of the human VIP gene 5' flanking sequence fused to a reporter. Subsequent gene deletion experiments revealed several regulatory regions on the gene, including a 645-bp sequence located approximately 4.0 upstream from the transcription start site. Here we examined this upstream region in detail. Inhibitory sequences were found to be present on each end of the 645-bp fragment. When removed, basal transcription increased more than 50-fold. Subsequent deletion/mutation analysis showed that the 213-bp fragment contained at least two enhancer elements. One of these was localized to an AT-rich 42-bp sequence shown by others to bind Oct proteins in neuroblastoma cells, while the other corresponded to a composite AP-1/ets element. In addition to these enhancers, a 28-bp sequence on the 213-bp fragment with no apparent homology to known silencers inhibited transcription. The studies provide molecular details of a complex regulatory region on the VIP gene that is likely to be used to finely tune the level of gene transcription in vivo.
Liu
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Department of Psychiatry and Mental Retardation Research Center, University of California at Los Angeles, 760 Westwood Plaza, Room 68-225 NPI, Los Angeles, California, 90024-1759
Krajniak
K
K
Chun
D
D
Sena
M
M
Casillas
R
R
Lelièvre
V
V
Nguyen
T
T
Bravo
D
D
Colburn
S
S
Waschek
J A
JA
D020121:5' Untranslated Regions / Q000235:genetics; D001483:Base Sequence; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D017930:Genes, Reporter; D051863:Host Cell Factor C1; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009447:Neuroblastoma / Q000378:metabolism; D050811:Octamer Transcription Factor-1; D050813:Octamer Transcription Factor-2; D012045:Regulatory Sequences, Nucleic Acid; D017384:Sequence Deletion; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D014660:Vasoactive Intestinal Peptide / Q000096:biosynthesis / Q000235:genetics*
D020121:5' Untranslated Regions; D004268:DNA-Binding Proteins; C497692:HCFC1 protein, human; D051863:Host Cell Factor C1; D050811:Octamer Transcription Factor-1; D050813:Octamer Transcription Factor-2; C494165:POU2F1 protein, human; C058435:POU2F2 protein, human; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D014660:Vasoactive Intestinal Peptide