Constitutive promoter occupancy by the MBF-1 activator and chromatin modification of the developmental regulated sea urchin alpha-H2A histone gene.
The tandemly repeated sea urchin alpha-histone genes are developmentally regulated. These genes are transcribed up to the early blastula stage and permanently silenced as the embryos approach gastrulation. As previously described, expression of the alpha-H2A gene depends on the binding of the MBF-1 activator to the 5' enhancer, while down-regulation relies on the functional interaction between the 3' sns 5 insulator and the GA repeats located upstream of the enhancer. As persistent MBF-1 binding and enhancer activity are detected in gastrula embryos, we have studied the molecular mechanisms that prevent the bound MBF-1 from trans-activating the H2A promoter at this stage of development. Here we used chromatin immunoprecipitation to demonstrate that MBF-1 occupies its site regardless of the transcriptional state of the H2A gene. In addition, we have mapped two nucleosomes specifically positioned on the enhancer and promoter regions of the repressed H2A gene. Interestingly, insertion of a 26 bp oligonucleotide between the enhancer and the TATA box, led to up-regulation of the H2A gene at gastrula stage, possibly by changing the position of the TATA nucleosome. Finally, we found association of histone de-acetylase and de-acetylation and methylation of K9 of histone H3 on the promoter and insulator of the repressed H2A chromatin. These data argue for a role of a defined positioned nucleosome in the promoter and histone tail post-translational modifications, in the 3' insulator and 5' regulatory regions, in the repression of the alpha-H2A gene despite the presence of the MBF-1 activator bound to the enhancer.
Di Caro
Valentina
V
Dipartimento di Biologia Cellulare e dello Sviluppo (Alberto Monroy), Università di Palermo, Parco d'Orleans II, 90128 Palermo, Italy.
Cavalieri
Vincenzo
V
Melfi
Raffaella
R
Spinelli
Giovanni
G
D000818:Animals; D020029:Base Pairing; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002843:Chromatin / Q000378:metabolism*; D015536:Down-Regulation; D004625:Embryo, Nonmammalian / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D005775:Gastrula / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D006655:Histone Deacetylases / Q000378:metabolism; D006657:Histones / Q000235:genetics*; D038101:Insulator Elements; D016254:Mutagenesis, Insertional; D009707:Nucleosomes / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D012097:Repressor Proteins / Q000378:metabolism; D015183:Restriction Mapping; D012617:Sea Urchins / Q000196:embryology* / Q000235:genetics; D015534:Trans-Activators / Q000378:metabolism*
D002148:Calmodulin-Binding Proteins; D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D012097:Repressor Proteins; D015534:Trans-Activators; D006655:Histone Deacetylases