ESTRADIOL DOWN REGULATES THE BINDING-ACTIVITY OF AN AVIAN VITELLOGENIN GENE REPRESSOR (MDBP-2) AND TRIGGERS A GRADUAL DEMETHYLATION OF THE MCPG PAIR OF ITS DNA-BINDING SITE

被引:46
作者
JOST, JP [1 ]
SALUZ, HP [1 ]
PAWLAK, A [1 ]
机构
[1] IRBM,ROME,ITALY
关键词
D O I
10.1093/nar/19.20.5771
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
A negative regulating protein (MDBP-2) from rooster liver nuclear extracts binds preferentially to a methylated promoter region 5'TTCACCTTmCGCTATGAGGGGGATCATACTGG3' of the avian vitellogenin II gene (Nucleic Acids Res. 19, 1029-1034, 1991). Treatment of adult and immature roosters with estradiol results in a 90% decrease in the binding activity of MDBP-2 within three days. This corresponds to the level found in egg laying hens. The decrease in the binding activity of MDBP-2 precedes the onset of vitellogenin gene transcription. At the same time, there is a two-fold increase in the binding activity of NHP-1 (tested with the same oligonucleotide as for MDBP-2), a protein thought to be involved in the active demethylation of DNA. The methylated oligonucleotide binds either MDBP-2 or NHP-1 and there is no complex formation between the two proteins and DNA. Estradiol treatment does not change the equilibrium binding constant of MDBP-2 which is about 10(-9)M for the methylated oligonucleotide. The early kinetics of demethylation of the mCpG pair in the binding site of MDBP-2 was studied by means of genomic sequencing. A low level of demethylation of mCpG starts gradually on both DNA strands already 4 hours after estradiol treatment during the lag phase of vitellogenin mRNA synthesis. It is concluded that the lowering of the binding activity of MDBP-2 may have a stronger effect on the derepression of the gene than the slow demethylation of MDBP-2 DNA binding site. The role of the methylated CpG is to assure a high binding affinity of the repressor to DNA.
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页码:5771 / 5775
页数:5
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