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Post by Max on Jul 17, 2005 6:43:31 GMT -5
Folate
Folate transport
Sp1
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Post by Max on Jul 17, 2005 6:46:24 GMT -5
Biochim Biophys Acta. 2005 Jan 21;1727(1):45-57. Epub 2004 Dec 31. Related Articles, Links
Transcriptional regulation of the human reduced folate carrier promoter C: synergistic transactivation by Sp1 and C/EBP beta and identification of a downstream repressor.
Payton SG, Whetstine JR, Ge Y, Matherly LH.
Department of Pharmacology, Wayne State University, School of Medicine, Detroit, MI, USA.
The human reduced folate carrier (hRFC) is ubiquitously but differentially expressed in human tissues and its levels are regulated by up to six alternatively spliced non-coding regions (designated A1/A2, A, B, C, D, and E) and by at least four promoters. By transient transfections of HepG2 human hepatoma cells with 5' and 3' deletion constructs spanning 2883 bp of upstream sequence, a transcriptionally important region was localized to within 177 bp flanking the transcriptional start sites for exon C. By gel shift and chromatin immunoprecipitation assays, Sp1 and C/EBP beta transcription factors were found to bind consensus elements (GC-box, CCAAT-box) within this region. The functional importance of these elements was confirmed by transient tranfections of HepG2 cells with hRFC-C reporter constructs in which these elements were mutated, and by co-transfections of Drosophila SL-2 cells with wild-type hRFC-C promoter and expression constructs for Sp1 and C/EBP beta. Whereas both Sp1 and C/EBP beta transactivated hRFC-C promoter activity, C/EBP alpha and gamma were transcriptionally inert. Sp1 combined with C/EBP beta resulted in a synergistic transactivation. In HepG2 cells, transfections with Sp1 and C/EBP beta both increased endogenous levels of hRFC-C transcripts. By 3' deletion analysis, a repressor sequence was localized to within 71 bp flanking the minimal promoter. On gel shifts, a novel transcriptional repressor was localized to within 30 bp. Collectively, these results identify transcriptionally important regions in the hRFC-C minimal promoter that include a GC-box and CCAAT-box, and suggest that cooperative interactions between Sp1 and C/EBP beta are essential for hRFC-C transactivation. Another possible factor in the tissue-specific regulation of the hRFC-C region involves the downstream repressor flanking the minimal promoter.
PMID: 15652157 [PubMed - indexed for MEDLINE]
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Post by Max on Jul 17, 2005 6:47:07 GMT -5
References:
Biochem J. 2004 Oct 15;383(Pt 2):249-57. Related Articles, Links
Roles of USF, Ikaros and Sp proteins in the transcriptional regulation of the human reduced folate carrier B promoter.
Liu M, Whetstine JR, Payton SG, Ge Y, Flatley RM, Matherly LH.
Experimental and Clinical Therapeutics Program, Barbara Ann Karmanos Cancer Institute, 110 E. Warren Ave., Detroit, MI 48201, USA.
The hRFC (human reduced folate carrier) is ubiquitously but differentially expressed in human tissues and its levels are regulated by up to seven non-coding regions (A1, A2, A, B, C, D and E) and at least four promoters. For the hRFC-B basal promoter, regulation involves binding of Sp (specificity protein) transcription factors to a critical GC-box. By transiently transfecting HT1080 cells with 5'- and 3'-deletion constructs spanning 1057 bp of upstream sequence, a transcriptionally important region was localized to 158 bp flanking the transcriptional start sites. By gel shift and chromatin immunoprecipitation assays, USF (upstream stimulatory factor), Sp1 and Ikaros-related proteins were bound to consensus elements (one E-box, two GC-box and three Ikaros) within this region. The functional importance of these elements was confirmed by transient tranfections of HT1080 cells with hRFC-B reporter constructs in which they were mutated, and by co-transfections of Drosophila Mel-2 cells with wild-type hRFC-B promoter and expression constructs for USF1, USF2a, Sp1 and Ikaros 2 and 8. Both USF1 and Sp1 proteins transactivated the hRFC-B promoter. Sp1 combined with USF1 resulted in a synergistic transactivation. Identical results were obtained with USF2a. Ikaros 2 was a repressor of hRFC-B promoter activity whose effects were partly reversed by the dominant-negative Ikaros 8. In HT1080 cells, transfection with Ikaros 2 decreased endogenous hRFC-B transcripts, whereas USF1 and Sp1 increased transcript levels. Ikaros 2 also decreased reporter gene activity and levels of acetylated chromatin associated with the endogenous promoter. Collectively, these results identify transcriptionally important regions in the hRFC-B promoter that include multiple GC-box, Ikaros and E-box elements. Our results also suggest that co-operative interactions between transcription factors Sp1 and USF are essential for high-level hRFC-B transactivation and imply that these effects are modulated by the family of Ikaros proteins and by histone acetylation.
PMID: 15214842 [PubMed - indexed for MEDLINE]
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Post by htryeeyue on Sept 7, 2018 19:31:21 GMT -5
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