Dpb11 bridges are induced by compromised DNA replication or sister chromatid decatenation. (A) Dpb11 bridges are induced by replication stress. Cells expressing Dpb11-YFP (ML533) were analyzed at the indicated time points after treatment with MMS (0.03%). For quantification, 300–700 cells were analyzed per time point. Bridges are defined as short bridges (<1 µm) or long bridges (>2 µm). (B) Dpb11 bridges form at the metaphase-to-anaphase transition (between 60 and 90 min after G1 release). Cells expressing Dpb11-YFP (ML533) were arrested in G1 phase with α-factor for 2.5 h, and subsequently released into fresh medium without α-factor. Pictures were taken at the indicated time points after G1 release. The length of bridges was measured as in A. (C) Quantitation of anaphase bridges after release from G1 arrest. For each time point in B, 150–350 cells were analyzed. Asterisk indicates a P value of <0.05 compared with time-point zero. (D) Dpb11 bridges accumulate in a top2-1 mutant. A temperature-sensitive top2-1 mutant (VS11-13D) and its corresponding TOP2 wild type (ML533) were examined for Dpb11-YFP bridges at the permissive temperature (25°C) or after incubation at the restrictive temperature (37°C) for 2 h. Indicated samples were treated with 0.03% MMS for 70 min. (E) Top2 localizes to UFBs. Cells expressing Top2-CFP (VS21) were stained with Hoechst and examined for Top2 bridges. (F) Top2 and Dpb11 colocalize on anaphase bridges. Cells expressing Dpb11-CFP and Top2-YFP (VS22-7B) were analyzed. (G) Dpb11 bridges accumulate in an smc6-9 mutant. A temperature-sensitive smc6-9 mutant (SMG266-6D) and its corresponding SMC6 wild type (ML533) were examined for Dpb11-YFP bridges after incubation at the restrictive (37°C) temperature for 2 h. Error bars represent 95% confidence intervals.