Abstract
The 5′-flanking and internal regions of the rabbit ot-globin gene, which constitute a CpG island, are required for enhancer-independent expression in transfected cells. In this study, electrophoretic mobility shift assays revealed that a battery of nuclear proteins from both erythroid and nonerythroid cells bind specifically to these regulatory regions. Assays based on exonuclease III digestion, methylation interference, and DNase I footprinting identified sequences bound by proteins in crude nuclear extracts and by purified transcription factor Sp1. In the 5′ flank, recognition sites for the transcription factors α-IRP (positions -53 to -44 relative to the cap site), CP1 (-73 to -69), and Sp1 (-95 to -90) are bound by proteins in K562 cell nuclear extracts, as are three extended upstream regions. Two recognition sites for Sp1 in intron 1 are also bound both by proteins in crude nuclear extracts and by purified Sp1. The sequences CCAC in intron 2 and C5 in the 3′-untranslated region also bind proteins. A major binding site found in exon 1, TATGGCGC, matches in sequence and methylation interference pattern the binding site for nuclear protein YY1, and binding is inhibited through competition by YY1-specific oligonucleotides. The protein-binding sites flanking and internal to the rabbit α-globin gene may form an extended promoter.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 5439-5449 |
| Number of pages | 11 |
| Journal | Molecular and Cellular Biology |
| Volume | 13 |
| Issue number | 9 |
| State | Published - Sep 1993 |
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All Science Journal Classification (ASJC) codes
- Cell Biology
- Genetics
- Molecular Biology
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Nuclear protein-binding sites in a transcriptional control region of the rabbit α-globin gene. / Yost, Susan E.; Shewchuk, Brian; Hardison, Ross Cameron.
In: Molecular and Cellular Biology, Vol. 13, No. 9, 09.1993, p. 5439-5449.Research output: Contribution to journal › Article
TY - JOUR
T1 - Nuclear protein-binding sites in a transcriptional control region of the rabbit α-globin gene
AU - Yost, Susan E.
AU - Shewchuk, Brian
AU - Hardison, Ross Cameron
PY - 1993/9
Y1 - 1993/9
N2 - The 5′-flanking and internal regions of the rabbit ot-globin gene, which constitute a CpG island, are required for enhancer-independent expression in transfected cells. In this study, electrophoretic mobility shift assays revealed that a battery of nuclear proteins from both erythroid and nonerythroid cells bind specifically to these regulatory regions. Assays based on exonuclease III digestion, methylation interference, and DNase I footprinting identified sequences bound by proteins in crude nuclear extracts and by purified transcription factor Sp1. In the 5′ flank, recognition sites for the transcription factors α-IRP (positions -53 to -44 relative to the cap site), CP1 (-73 to -69), and Sp1 (-95 to -90) are bound by proteins in K562 cell nuclear extracts, as are three extended upstream regions. Two recognition sites for Sp1 in intron 1 are also bound both by proteins in crude nuclear extracts and by purified Sp1. The sequences CCAC in intron 2 and C5 in the 3′-untranslated region also bind proteins. A major binding site found in exon 1, TATGGCGC, matches in sequence and methylation interference pattern the binding site for nuclear protein YY1, and binding is inhibited through competition by YY1-specific oligonucleotides. The protein-binding sites flanking and internal to the rabbit α-globin gene may form an extended promoter.
AB - The 5′-flanking and internal regions of the rabbit ot-globin gene, which constitute a CpG island, are required for enhancer-independent expression in transfected cells. In this study, electrophoretic mobility shift assays revealed that a battery of nuclear proteins from both erythroid and nonerythroid cells bind specifically to these regulatory regions. Assays based on exonuclease III digestion, methylation interference, and DNase I footprinting identified sequences bound by proteins in crude nuclear extracts and by purified transcription factor Sp1. In the 5′ flank, recognition sites for the transcription factors α-IRP (positions -53 to -44 relative to the cap site), CP1 (-73 to -69), and Sp1 (-95 to -90) are bound by proteins in K562 cell nuclear extracts, as are three extended upstream regions. Two recognition sites for Sp1 in intron 1 are also bound both by proteins in crude nuclear extracts and by purified Sp1. The sequences CCAC in intron 2 and C5 in the 3′-untranslated region also bind proteins. A major binding site found in exon 1, TATGGCGC, matches in sequence and methylation interference pattern the binding site for nuclear protein YY1, and binding is inhibited through competition by YY1-specific oligonucleotides. The protein-binding sites flanking and internal to the rabbit α-globin gene may form an extended promoter.
UR - http://www.scopus.com/inward/record.url?scp=0027304447&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0027304447&partnerID=8YFLogxK
M3 - Article
C2 - 8355692
AN - SCOPUS:0027304447
VL - 13
SP - 5439
EP - 5449
JO - Molecular and Cellular Biology
JF - Molecular and Cellular Biology
SN - 0270-7306
IS - 9
ER -