High-fidelity chromosomal segregation requires the properly timed establishment of sisterchromatid cohesion mediated by the Cohesin complex, and its resolution at the metaphaseto-anaphase transition. We have examined cell-cycle progression in a yeast strain from which the origin recognition complex protein Orc2 was depleted after the assembly of prereplication complexes. We find that Orc2 depletion causes a delay in progression through mitosis, reflecting activation of both the DNAdamage and Mad2-spindle checkpoints. Surprisingly, sister-chromatid cohesion is impaired in Orc2-depleted cells, although Cohesin subunits are properly associated with chromatin. Reexpression of Orc2 in late G2/M phase restores chromatid cohesion. Finally, the targeting of Orc2 to a specific chromosomal locus suppresses premature sister-chromatid separation locally in a temperature-sensitive cohesin mutant. We conclude that ORG mediates sisterchromatid interaction on a pathway that is additive with Cohesin-mediated pairing.
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