Spo11 Function in Yeast Meiosis

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Meiotic recombination is initiated by DNA double-strand breaks (DSBs). The meiotic recombination pathway is understood best in budding yeast: An intact DNA duplex is cleaved by Spo11 protein to yield a covalent Spo11-DNA complex. Spo11 is then released and the 5'-terminal strands are degraded to yield 3'-single-stranded tails. These tails undergo strand invasion into an intact, homologous duplex, giving rise ultimately to mature products in either crossover or noncrossover configuration.

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After a DSB is made, Spo11 protein must be removed so that the DSB can be repaired, but the mechanism for removal was not clear. We recently demonstrated that Spo11 is removed by single-stranded endonucleolytic cleavage releasing Spo11 covalently attached to a short oligonucleotide (Neale et al., 2005). We also showed that this mechanism is conserved by demonstrating the presence of SPO11-oligo complexes in extracts from mouse testis. Surprisingly, we observed two discrete species of Spo11-oligo complex that differed with respect to the length of the attached oligonucleotide. We proposed that Spo11 is released by a pair of asymmetrically placed nicks. Such a scenario would have interesting implications for understanding how later asymmetry in the repair of DSBs might be accomplished.

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Spo11 is the catalytic center of the meiotic DSB-forming machinery, but it does not act alone. In yeast, at least nine other proteins are required for Spo11-dependent DSB formation. However, very little is known about their functions. To address this issue, we analyzed interactions involving Spo11 and the other DSB proteins (Arora et al., 2004), defining an interaction network that connects all of the DSB proteins together. This analysis identified Ski8 as a likely direct partner of Spo11. The connection to DSB formation had long been puzzling for Ski8, because the protein has direct roles in RNA metabolism in the cytoplasm. We demonstrated that Ski8 functions in RNA metabolism are separate from its requirement in DSB formation (Arora et al., 2004). Furthermore, we showed that Ski8 relocalizes from the cytoplasm to the nucleus during meiosis, strictly dependent on interaction between Ski8 and Spo11.

[Enlarge Image] Association of Rec102 with Meiotic Chromosomes

We also examined genetic and physical interactions between the Rec102 and Rec104 proteins, which are mutually dependent for proper sub-cellular localization, and which share a requirement for Spo11 and Ski8 for their recruitment to chromosomes (Kee et al., 2004). We explored the interaction of both proteins with meiotic chromosomes. Our findings support a model in which Rec102/Rec104 are involved in the chromatin loop-chromosome axis interactions known to be important for meiotic recombination.

DSB formation is limited to a narrow window of time during meiotic prophase, but what controls this timing is not well understood. We took a step in addressing this issue by showing that cyclin-dependent kinase (CDK) directly promotes DSB formation (Henderson et al., 2006). Specifically, we showed that CDK directly phosphorylates Mer2 (another protein required for DSB formation) and thereby controls the ability of Mer2 to promote DSB formation by controlling the interaction of Mer2 with other DSB proteins. These results suggest that regulation of DSB formation by CDK is part of the mechanism that coordinates recombination with progression through meiotic prophase.

Crossover formation is controlled such that each chromosome gets at least one crossover despite a low average number of crossovers per chromosome, and multiple crossovers on the same chromosome tend to be evenly and widely spaced. More DSBs are formed than crossovers and DSBs tend to be randomly placed relative to one another, so crossover control involves a decision by which a subset of DSBs enters a pathway that culminates in crossover formation, while all other DSBs follow a pathway(s) that generates primarily noncrossover products. To understand the logic of this decision, we examined recombination when breaks are reduced through the use of a series of yeast partial loss-of-function spo11 mutants (Martini et al., 2006). We found that crossovers tend to be maintained at the expense of noncrossovers, a phenomenon we refer to as "crossover homeostasis." These findings defined a previously unsuspected manifestation of crossover control, i.e., that the crossover/noncrossover ratio can change to maintain crossovers. Our results support the hypothesis that an obligate crossover is a genetically programmed event tied to crossover interference.