In vivo and in vitro protein-DNA interactions at the distal oestrogen response element of the chicken vitellogenin gene: evidence for the same protein binding to this sequence in hen and rooster liver.
The major egg white protein, vitellogenin, is synthesized in a tissue specific and oestradiol dependent manner in the liver of egg-laying hens. In this paper, we describe a detailed study of the protein-DNA interactions at the distal oestrogen response element (ERED) located 600 bp upstream of the start of transcription. In vivo footprinting of hepatocytes from adult hens and roosters with 0.5-0.0005% dimethylsulphate (DMS) revealed, at critical concentrations of DMS, protection of distinct guanosine residues within the ERED and adjacent downstream sequence in both cases. From this, it was concluded that there were proteins present in both tissues binding to this region in vivo. In vitro studies using missing base contact probing and proteolytic clipping band shift assays with hen and rooster liver nuclear extracts identified the ERE binding protein to be the same or very closely related in both tissues. Furthermore, the protein from rooster nuclear extracts bound to the ERE sequence even when the DNA was methylated at CpG dinucleotides, u.v. cross-linking experiments performed with bromodeoxyuridine substituted ERE, revealed that a nuclear protein with Mr of about 75,000-80,000 bound specifically to this sequence. These studies demonstrate that apart from the oestrogen receptor, at least one other protein can interact specifically with the chicken vitellogenin ERE, independently of hormonal expression of the gene.
McEwan
I J
IJ
Friedrich Miescher-Institut, Basel, Switzerland.
Saluz
H P
HP
Jost
J P
JP
D000818:Animals; D001483:Base Sequence; D002645:Chickens; D003432:Cross-Linking Reagents; D004247:DNA / Q000378:metabolism / Q000528:radiation effects; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004967:Estrogens / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006868:Hydrolysis; D008099:Liver / Q000378:metabolism; D008297:Male; D008745:Methylation; D008969:Molecular Sequence Data; D011687:Purines / Q000378:metabolism; D011743:Pyrimidines / Q000378:metabolism; D014819:Vitellogenins / Q000235:genetics*
D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D004967:Estrogens; D011687:Purines; D011743:Pyrimidines; D014819:Vitellogenins; D004247:DNA