The control of Spo11's interaction with meiotic recombination hotspots.
Programmed double-strand breaks (DSBs), which initiate meiotic recombination, arise through the activity of the evolutionary conserved topoisomerase homolog Spo11. Spo11 is believed to catalyze the DNA cleavage reaction in the initial step of DSB formation, while at least a further 11 factors assist in Saccharomyces cerevisiae. Using chromatin-immunoprecipitation (ChIP), we detected the transient, noncovalent association of Spo11 with meiotic hotspots in wild-type cells. The establishment of this association requires Rec102, Rec104, and Rec114, while the timely removal of Spo11 from chromatin depends on several factors, including Mei4 and Ndt80. In addition, at least one further component, namely, Red1, is responsible for locally restricting Spo11's interaction to the core region of the hotspot. In chromosome spreads, we observed meiosis-specific Spo11-Myc foci, independent of DSB formation, from leptotene until pachytene. In both rad50S and com1Delta/sae2Delta mutants, we observed a novel reaction intermediate between Spo11 and hotspots, which leads to the detection of full-length hotspot DNA by ChIP in the absence of artificial cross-linking. Although this DNA does not contain a break, its recovery requires Spo11's catalytic residue Y135. We propose that detection of uncross-linked full-length hotspot DNA is only possible during the reversible stage of the Spo11 cleavage reaction, in which rad50S and com1Delta/sae2Delta mutants transiently arrest.
Prieler
Silvia
S
Institute of Botany, Max F. Perutz Laboratories, Department of Chromosome Biology, A-1030 Vienna, Austria.
Penkner
Alexandra
A
Borde
Valérie
V
Klein
Franz
F
D019943:Amino Acid Substitution / Q000235:genetics; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D019457:Chromosome Breakage / Q000235:genetics / Q000502:physiology; D015825:Chromosomes, Fungal / Q000235:genetics* / Q000502:physiology; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D004706:Endodeoxyribonucleases; D004950:Esterases / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal / Q000235:genetics / Q000502:physiology; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D049471:Pachytene Stage / Q000235:genetics* / Q000502:physiology; D017354:Point Mutation / Q000235:genetics; D011995:Recombination, Genetic / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D013573:Synaptonemal Complex / Q000235:genetics* / Q000378:metabolism; D000099257:Meiotic Recombination Protein SPO11
D002843:Chromatin; D004271:DNA, Fungal; D004706:Endodeoxyribonucleases; D004950:Esterases; D046912:Multiprotein Complexes; D029701:Saccharomyces cerevisiae Proteins; D000099257:Meiotic Recombination Protein SPO11