Bloom’s syndrome is a rare autosomal recessive disorder caused by defects in BLM, a RecQ helicase involved in DNA repair. Bloom’s patients exhibit immunodeficiency and tumour predisposition with a 90% chance of developing cancer by age 25. At a cellular level, Bloom’s patient cell lines display genomic instability characterized by increased frequency of sister chromatid exchanges and excessive homologous recombination.  Since cancer risk is sporadic, and not specific to any particular type, BLM is essential in all tissues for tumor suppression and genome stability.  

When purified in isolation, the BLM protein has been shown to unwind double stranded DNA associated with breaks that occur during recombination and replication. However, BLM exists in a nuclear protein containing Topoisomerase IIIα, a type 1A topoisomerase, RMI1 (Replication-Mediated Instability 1) and RMI2.  Topoisomerase IIIα relieves negatively supercoiled DNA, RMI1 stimulates topoisomerase IIIα activity, and RMI2 stabilizes the complex.  In our laboratory, all four human proteins have been expressed simultaneously in insect cells using the Multibac system. We performed functional and structural studies to determine how BLM interacts with DNA and other members of the purified complex.  Gel shift assays demonstrate that BLM plays an important role in targeting the complex to DNA substrates that contain single strand-double strand DNA junctions such as 3’- or 5’-flaps.  In addition, pulldown experiments with flag tagged fragments show that BLM may interact with the zinc finger domains of Topoisomerase IIIα.

Using electron microscopy we have determined the overall architecture of the Bloom’s complex, and a model indicating its critical role in DNA repair will be presented.