Multisubunit erythroid complexes binding to the enhancer element of the chicken histone H5 gene.
We identified the factor(s) that bind to the chicken erythroid-cell-specific histone H5 enhancer region which is located on the 3' end of the gene. In DNAase I footprinting and u.v. cross-linking experiments with nuclear extracts from adult chicken immature erythrocytes, we determined that the trans-acting factor GATA-1 was the predominating protein interacting with the histone H5 enhancer. GATA-2 and GATA-3 were not detected. In contrast, gel-mobility-shift assays and competition experiments demonstrated that several specific complexes formed with the histone H5 enhancer region. Gel-mobility-shift assays with 23 bp oligonucleotides containing the GATA-binding site (AGATAA) of the histone H5 enhancer or of the beta-globin enhancer showed that the GATA sequence was sufficient for the formation of at least five complexes. Diagonal mobility-shift assays demonstrated that multisubunit complexes were forming with the GATA-1 protein. Our interpretation of the results is that GATA-1 interacts with a protein of approx. 105 kDa which, in turn, can associate with protein or protein complexes of approx. 26 kDa, 146 kDa and a protein(s) of molecular mass greater than 450 kDa. The different multisubunit complexes formed via the trans-acting factor GATA-1 may impart different transcriptional responses to the promoter and enhancer elements of the histone H5 and globin genes.
Penner
C G
CG
Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Manitoba, Winnipeg, Canada.
Davie
J R
JR
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000737:chemistry; D002645:Chickens; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I; D004742:Enhancer Elements, Genetic; D004912:Erythrocytes / Q000737:chemistry* / Q000648:ultrastructure; D050981:Erythroid-Specific DNA-Binding Factors; D005914:Globins / Q000235:genetics; D006657:Histones / Q000235:genetics*; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014466:Ultraviolet Rays; D016335:Zinc Fingers
D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D006657:Histones; D009841:Oligonucleotides; D014157:Transcription Factors; D005914:Globins; D004247:DNA; D003850:Deoxyribonuclease I