Transcriptional activity of the SHP-1 gene in MCF7 cells is differentially regulated by binding of NF-Y factor to two distinct CCAAT-elements.
Our previous studies have shown that SHP-1, a SH2 domain-containing protein-tyrosine phosphatase, is expressed not only in cells of hematopoietic lineages, but also in many non-hematopoietic cells under the control of an alternative tissue-specific promoter, P1. In this study, the activity of the P1 promoter was analyzed in a region spanning 3.5 kb upstream of the major transcription start site in non-hematopoietic MCF-7 cells. Using DNA footprinting, gel retardation assays and mutational analysis, we have characterized cis-regulatory elements that are essential to confer the P1 promoter activity. An upstream Sp1 element (-126 to -118) positively regulated this TATA-box-lacking promoter. Two inverted CCAAT-elements (-332 to -328 and -66 to -62) played important roles in regulating the SHP-1 gene expression, and transcription factor NF-Y predominantly bound to the two CCAAT-elements. Binding of NF-Y to the distal CCAAT-element enhanced the transcriptional activity of the P1 promoter. In contrast, binding of NF-Y to the proximal CCAAT-element and interacting with repressor(s) inhibited the promoter activity. Furthermore, incubation of MCF7 cells with 100 ng/ml trichostatin A, an inhibitor of histone deacetylase, significantly increased the activity of the P1 promoter. Mutation in the proximal CCAAT-element, however, eliminated the activating effect of trichostatin A on the promoter. Together, our data suggest that NF-Y factor can function either as a specific positive or negative regulator of P1 promoter activity in non-hematopoietic MCF7 cells.
Xu
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Department of Animal Science, Macdonald Campus, McGill University, Ste. Anne de Bellevue, H9X 3V9, Quebec, Canada.
Banville
D
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Zhao
H F
HF
Zhao
X
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Shen
S H
SH
D001483:Base Sequence; D001665:Binding Sites; D023081:CCAAT-Binding Factor / Q000378:metabolism*; D018076:DNA, Complementary; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000235:genetics*; D016329:Sp1 Transcription Factor / Q000378:metabolism; D016385:TATA Box; D014157:Transcription Factors; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D023081:CCAAT-Binding Factor; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C492061:NFYA protein, human; C492063:NFYB protein, human; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; C509249:PTPN6 protein, human; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases