Nuclear protein-binding sites in a transcriptional control region of the rabbit α-globin gene

Susan E. Yost, Brian Shewchuk, Ross Cameron Hardison

Research output: Contribution to journalArticle

19 Citations (Scopus)

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 languageEnglish (US)
Pages (from-to)5439-5449
Number of pages11
JournalMolecular and Cellular Biology
Volume13
Issue number9
StatePublished - Sep 1993

Fingerprint

Globins
Nuclear Proteins
Protein Binding
Binding Sites
Rabbits
Complex Mixtures
Introns
Methylation
Genes
Proteins
Sp1 Transcription Factor
Erythroid Cells
CpG Islands
K562 Cells
5' Flanking Region
Nucleic Acid Regulatory Sequences
Deoxyribonuclease I
3' Untranslated Regions
Electrophoretic Mobility Shift Assay
Cell Extracts

All Science Journal Classification (ASJC) codes

  • Cell Biology
  • Genetics
  • Molecular Biology

Cite this

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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.",
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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 journalArticle

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