Journal
GENETICS
Volume 169, Issue 4, Pages 1973-1983Publisher
GENETICS SOCIETY AMERICA
DOI: 10.1534/genetics.104.039230
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Funding
- NIGMS NIH HHS [R01 GM031693, F32 GM020190, R56 GM031693, GM20190, GM31693] Funding Source: Medline
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The ade6-M26 mutation of Schizosaccharomyces pombe created a meiotic recombination hotspot. Previous analyses indicated that the heptamer 5'-ATGACGT-3' was necessary and sufficient for hotspot activity; the Atf1-Pcr1 transcription factor binds to this sequence and activates M26. After finding cases in which the M26 heptamer in ade6 was, surprisingly, not active as a hotspot, we used an in vitro selection method (SELEX) that revealed an 18-bp consensus sequence for Atf1-Pcr1 binding, 5'-GNVTATGACGTCATNBNC-3', containing the M26 heptamer at its core. Using this consensus sequence as a guide, we made mutations on each side of the heptamer at two separate sites in ade6. These mutations increased the intracellular hotspot activity of the heptamer, in some cases by > 15-fold. These results show that M26, the eukaryotic recombination hotspot with the most precisely defined nucleotide sequence, is larger than previously thought, and they provide valuable information for clarifying the role of M26, and perhaps other hotspots, in meiotic recombination.
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