The Contribution of Genotoxic Stress to the Formation of Chromosome Bridges in Anaphase

Defects in enzymes (e.g., Blooms syndrome helicase (BLM)) that unwind and detangle DNA during replication are associated with genomic instability and human disease. Interestingly, BLM localizes to late segregating chromosome elements called chromosome bridges that are likely to be tangled sister chromatids that arise during replication. However, it is unknown whether genotoxic stresses promote the formation of chromosome bridges or inhibit their resolution during anaphase. To address this question, we propose to challenge budding yeast cells expressing histone H1 fused to GFP with the following compounds: hydroxyurea, an inhibitor of DNA replication; MMS, induces single strand lesions; razoxane, aclurubucin and camptothecin,  inhibitors of topoisomerases, enzymes important for the resolution of tangled sister chromatids. We have previously observed that inhibition of DNA replication following HU treatment gives rise to slower to resolve chromosome bridges. Preliminary observations indicate that MMS treatment does not affect chromosome bridge formation or resolution. Further experiments are in progress to further characterize the HU-induced chromosome bridges and to evaluate the effect of other genotoxic stresses on chromosome bridges. The long-term goal is to link the behavior of chromosome bridges to the severity of genomic instability by monitoring DNA double-strand breaks. This may have important implications for cancer onset and therapeutic strategies.