Locally Unwound DNA

The high AT content is a feature of DNA sequences that, under superhelical tension, may form unpaired structures, termed DNA unwinding elements (DUE) or base unpairing regions (BUR). Such sequences are a common feature of bacterial and yeast replication origins and are associated with some human replication origins. As part of the pentanucleotide expansion project (see below) we studied the formation of unpaired structures in several plasmids containing human genomic sequences composed of (ATTCT)•(AGAAT) repeats together with their AT-rich flanks. At moderate superhelical densities, DNA unwinding in the (ATTCT)•(AGAAT) repeats is detected by several methods. At higher superhelicity DNA unwinding extends into adjacent flanks of slightly lesser AT percentage. The superhelical energy required to initiate duplex unpairing is essentially length independent at more than 8 repeats. In plasmids with 5 and less repeats superhelical tension induces base unpairing in other available AT-rich sequences. When the size of an unwound region is large enough, unpaired DNA strands condense into globular structures. The stably unpaired DNA structure has a potential to serve as an aberrant replication origin because it supported complete plasmid replication in a HeLa cell extract in the absence of a replication initiation protein [1]. Locally unpaired DNA may also serve as a binding site for poly(ADP-ribose) polymerase (PARP-1) which recognizes single- and double-strand breaks in DNA and a number of structural distortions in unbroken DNA [2]. PARP-1 binding to unpaired DNA activates the enzyme for auto- and trans-modification of nuclear proteins, and may therefore, be related to the PARP-1's roles in transcription, replication and recombination.

Publications

1. Potaman VN, Bissler JJ, Hashem VI, Oussatcheva EA, Lu L, Shlyakhtenko LS, Lyubchenko YL, Matsuura T, Ashizawa T, Leffak M, Benham CJ, Sinden RR. (2003) Unpaired structures in SCA10 (ATTCT)n•(AGAAT)n repeats. J. Mol. Biol. 326: 1095-1111 (request a copy).
2. Lonskaya I, Potaman VN, Shlyakhtenko LS, Oussatcheva EA, Lyubchenko YL, Soldatenkov VA. (2005) Regulation of poly(ADP-ribose) polymerase-1 by DNA structure-specific binding. J. Biol. Chem. 280, 17076-17083 (link to PDF)

Supercoil-induced unwinding of DNA strands in plasmid DNA. The unpaired "bubble" maps to DNA sequence containing 29 (ATTCT)•(AGAAT) repeats (80% AT-rich) and 80 base pairs of an adjacent mixed sequence (75% AT-rich).