Sense of superhelical writhe of a bent DNA influences eukaryotic promoter function

Curved DNA structures are sometimes reported to reside within the transcriptionaJ control regions of prokaryotic and cukaryotir genes. Extensive data -iiggestiiig a roi(i in prokaryotic transcription for tiie structure has accumulated over the last few years. However, little has been revealed as to the role of DNA curvature in eukaryotic transcription. We investigated the effects on transcription of DNA curvatures located in the upstream region of an eukary otic promoter. Based on the knowledge from the study of Calladirte et ai ( 1988). double-stranded oligormcleotides having various three-dimensional architectures were designed, synthesized and ligated to the herpes simplex virus thymidine kinase ( TK) promoter. Furthermore, rotational orientations of these oligonudeotides relative to the promoter were altered by deleting several base pairs from the region located between each synthetic DNA segment and the promoter. The effects of DNA curvatures on the promoter strength were investigated by employing tuciferase assay. It was revealed that the DNA segment having a leftward writhe greatly influenced the strength of the TA'promoter in a rotational orientation dependent manner. In vitro nucleosome reconstitution experiments were also carried out by employing HeLa core histones. The result suggested that the promoters having a leftward writhe in their upstreams were favored for incorporation into nucleosomes. Possible mechanistic roles of DNA curvatures in eukaryotic Iranscription will be di-4russed.