DNA Unwinding in Control Regions of Yeast and E. coli.
The formation of an unwound DNA region may well be an essential step in gene transcription. The size and state of such unwound regions is yet far from clear. Currently available tools to study DNA unwinding include: a. Single strand specific nucleases to identify unwound regions. b. Two dimensional topoisomer analysis to determine extent of unwinding. c. Conformation specific DNA reagents which can distinguish between the various paranemic structures unwound DNA can assume (cf. Yagil, Crit. Revs. BMB. 26: 475, 1991). Purine.pyrimidine (R.Y) tracts are highly overrepresented in yeast promoter regions (Yagil, Yeast 10: 603). The three techniques were consequently applied to two strong yeast promoter regions DED1 and CYC1, that contain long R.Y tracts.
The principal P1 cleavage points of the DED1 promoter region map within a sequence containing a pyrimidine tract of 40 bases. In the promoter of CYC1, a region of 33/36 pyrimidines is the principal P1 sensitive region (cf. Yagil et al., Gene, 225: 152 1998). 2d-topoisomer analysis indicates the unwinding of 6 primary turns. The limited symmetry of the cleaved polypyrimidine regions favors the formation of a paranemic duplex in the unwound region. Previous work showed, by 2d -topoisomer analysis, that the chicken beta globin promoter and SV40 control regions also assume an unwound state. It is proposed that polypyrimidine tracts, in addition to A,T rich tracts, serve as DNA unwinding centers (DUE) in eukaryotic genes. In prokaryotic genes, the A,T binary DNA motif was found to prevail (Shomer and Yagil, NAR 27: 4491, 1999). In each case, the length of the unwound region could determine the number of polymerase complexes entering a gene, and thus control the frequency of gene transcription.
Dept. of Molecular Cell Biology, the Weizmann Institute