SUNY at Albany
June 19-23, 2001
Binary DNA Tracts can serve as DNA Unwinding Centers.
Binary DNA tracts are DNA tracts composed of only two bases on any one of the complementary strands. Four combinations exist: R.Y: Purines (A,G) on one strand and pyrimidines (C,T) on the complimentary one; K.M: (G,T) on one strand and (A,C) on the other; W: (A,T) and S: (G,C), auto complementary between the strands.
We and others detected a massive over representation of long binary DNA tracts(10-100 nt) in all three kingdoms of life. In most eukaryotes examined so far, the most highly over represented tracts are of the R.Y motif, as established experimentally by Chargaff and coworkers already in the 1950's. The high extent of over representation is typical for genomes from yeast to humans. The over representations are highest in intergenic regions as compared with coding regions, with introns occupying an intermediate position. The highest excess is observed in promoter regions. Thus, a 46 fold higher concentration of R.Y tracts longer than 15 nt, compared with randomized DNA of the same composition, is found in the 183 promoter regions of yeast chromosome III (Yagil, Yeast, 10: 603-611, 1994). More recently we examined chromosome 2 of Arabidopsis (19.6 Mb). This chromosome manifests an excess of 47.5 fold of R.Y tracts ³15 in its promoters!. In prokaryotes (E. coli, H. influenzae) however , the W tracts are the most highly over represented theme (Yagil, Nucleic Acid Res., 27: 4491-4500, 1999).
We have now studied the first annotated sequence of D. melongaster, the 3 Mb Adh-cactus region on chr. 2L. Surprisingly, the most over represented binary motif in this region are the W tracts, rather than the R.Y tracts in other eukaryotic chromosomes examined so far. Thus, Adh-cactus promoter regions (400 bases upstream) have 60,519 bases in W tracts of ³15, compared with 15,058 bases in R.Y promoter tracts ³15. This is an 13.65 fold over representation compared with randomized DNA of the same composition. The Adh cactus region thus resembles E. Coli promoters, which have 17.6 times more W tracts than random DNA. The relative ease of melting of the A,T rich tracts suggests that the excessive W tracts have a role in DNA strand separation, serving as DNA unwinding centers. This idea is supported by recent experimental studies of E. coli promoters by Grouse, Sheridan and coworkers ("UP elements").
We have tested experimentally whether an unwinding role can be assigned also to the R.Y tracts in the eukaryotic promoters. Two yeast promoters containing a long poly-pyrimidine tract on the coding strand (32 nt on the Cyc1 promoter, 39 nt on the Ded1 promoter) were subjected to single strand specific nucleases (Yagil, Shimron and Tal, Gene, 225: 152-163, 1998). The polypyrimidine tracts were found to be preferentially digested by P1 nuclease when in a negatively supercoiled plasmid. Two dimensional topoisomer analysis showed that up to six turns of the helix are opened, i.e., some 60 DNA bases become unwound at normal cellular superhelicities. Based on these data and on similar data in other species, it is proposed that the excessive binary DNA sequences do serve as DNA unwinding centers, termed DUE's by Kowalski and coworkers. The DUE's in the promoters can be expected to act as controlling transcriptional frequencies by controlling the rate of polymerase entrances into gene promoters. In support of this notion we consistently observe that the longer binaries reside in promoters of genes with high transcription frequencies.
Dept. of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot, Israel, 76100; Tel. 972-89-460-918; Fax 972-89-344-125. E-mail: email@example.com; Web page: www.weizmann.ac.il/~lcyagil.