We identify Xenopus NF-Y as a key regulator of acetylation responsiveness for the Xenopus hsp70 promoter within chromatin assembled in Xenopus oocyte nuclei. Y-box sequences are required for the assembly of DNase I-hypersensitive sites in the hsp70 promoter, and for transcriptional activation both by inhibitors of histone deacetylase and by the p300 acetyltransferase. The viral oncoprotein E1A interferes with both of these activation steps. We clone Xenopus NF-YA, NF-YB and NF-YC and establish that NF-Y is the predominant Y-box-binding protein in Xenopus oocyte nuclei. NF-Y interacts with p300 in vivo and is itself a target for acetylation by p300. Transcription from the hsp70 promoter in chromatin can be enhanced further by heat shock factor. We suggest two steps in chromatin modification at the Xenopus hsp70 promoter: first the binding of NF-Y to the Y-boxes to preset chromatin and second the recruitment of p300 to modulate transcriptional activity.

We have cloned a member of the nuclear receptor superfamily. The cDNA was isolated from a rat liver library and encodes a protein of 446 aa with a predicted mass of 50 kDa. This clone (OR-1) shows no striking homology to any known member of the steroid/thyroid hormone receptor superfamily. The most related receptor is the ecdysone receptor and the highest homologies represent <10%v in the amino-terminal domain, between 15-37% in the carboxyl-terminal domain and 50-62% in the DNA binding domain. The expression of OR-1 appears to be widespread in both fetal and adult rat tissues. Potential DNA response elements composed of a direct repeat of the hexameric motifAGGTCA spaced by 0-6 ntwere tested in gel shift experiments. OR-1 was shown to interact with the 9-cisretinoic acid receptor (retinoid X receptor, RXR) and the OR-

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Gene expression requires binding of transcription factors to their cognate DNA response elements, the latter being often integrated into sequence-specifically positioned nucleosomes. To investigate the constraints imposed on factor-DNA recognition by the nucleosomal organization, we studied the binding of glucocorticoid receptor to a single glucocorticoid response element (GRE) displaying four different rotational frames in three different translational positions in reconstituted nucleosomes. We demonstrate that rotational setting of the GRE per se is important for its accessibility. Furthermore, the effects of rotational positioning of the GRE are different for different translational positions of the GRE in the nucleosome. A GRE placed near the nucleosomal dyad is totally blocked by rotating it 180 degrees so that the major groove of the GRE faces the histone octamer. If, on the other hand, the GRE is placed about 40 bp from the nucleosome dyad, then the 180 degrees rotation of the GRE still allows glucocorticoid receptor binding, albeit with a sixfold lower affinity than the peripherally oriented GRE. This suggests that both the rotational positioning and the translational positioning function as a framework for transcription factor response elements in gene regulation.