Pds5 is required for homologue pairing and limits chromosome compaction during meiosis. (A) Monitoring homologue pairing and sister chromatid cohesion during meiosis. CEN5 on both chromosome V homologues was marked by tetO/tetR-GFP and visualized by fluorescence microscopy. Wild-type and P_CLB2PDS5 cells were arrested at pachytene with ndt80Δ. Four cell types were observed in P_CLB2PDS5 cells: type I, a single GFP spot, indicating that homologues are paired; type II, two GFP spots, indicating that homologues fail to pair but sister chromatids stay together; type III, three GFP spots, indicating that homologues fail to pair and one pair of sister chromatids separate; and type IV, four GFP spots, indicating that homologues fail to pair and sister chromatid cohesion is lost on both homologues. (B) Quantitative measurement of homologue pairing and sister chromatid cohesion. At least 200 cells were scored for each strain. (C) Measurement of the axial length of chromosome V. Meiotic nuclear spreads were prepared. Chromosome V was identified by CEN5-GFP signal, and the entire length of the chromosome was determined by measurement of the Rec8-3HA staining (detected as in A). (D) The long arm of chromosome IV was marked by GFP at two loci (CEN4 and TEL4) with the lacO/lacI-GFP system. Only one homologue of chromosome IV was marked in these cells. (E) The length of chromosome IV arm was determined by measurement of the distance between two GFP spots. Cells were induced to enter synchronous meiosis, and aliquots were withdrawn at the indicated times for preparation of immunofluorescence. Error bars indicate SD. (F) Representative images from three time points are shown. WT, wild type.