DNA unwinding in the CYC1 and DED1 yeast promoters

The capacity of promoter DNA of two yeast genes to be unwound was studied. Both promoters, those of the CPC1 and DED1 genes, contain long oligopurine. oligopyrimidine (R.Y) tracts. The two promoters were cloned into negatively supercoiled plasmids, and their sensitivity to single-strand specific nucl ease pi was examined. Extensive P1 cleavage was located within the R.Y trac ts, and cleavage sites were mapped. The extent of cleavage was only slightl y dependent on P1 concentration, indicating a slow conversion of an interme diate form of DNA into the P1 reactive state. The cleavage required negativ e supercoiling and was suppressed by NaCl, MgCl2 and spermine. Two-dimensio nal topoisomer analysis showed that six superhelical turns were opened in t he plasmids examined. The results indicate that at sufficient torsional str ess, the R.Y tracts can intermittently undergo a transition into an unwound , ready-to-separate state. The oligopurine.oligopyrimidine tracts may thus serve as DNA unwinding centers in the gene promoters where they reside. (C) 1998 Elsevier Science B.V. All rights reserved.