A Multiprotein Nuclear Complex Connects Fanconi Anemia and Bloom Syndrome

Abstract

Bloom syndrome (BS) is a genetic disorder associated with dwarfism, immunodeficiency, reduced fertility, and an elevated risk of cancer. To investigate the mechanism of this disease, we isolated from human HeLa extracts three complexes containing the helicase defective in BS, BLM. Interestingly, one of the complexes, termed BRAFT, also contains five of the Fanconi anemia (FA) complementation group proteins (FA proteins). FA resembles BS in genomic instability and cancer predisposition, but most of its gene products have no known biochemical activity, and the molecular pathogenesis of the disease is poorly understood. BRAFT displays a DNA-unwinding activity, which requires the presence of BLM because complexes isolated from BLM-deficient cells lack such an activity. The complex also contains topoisomerase III␣ and replication protein A, proteins that are known to interact with BLM and could facilitate unwinding of DNA. We show that BLM complexes isolated from an FA cell line have a lower molecular mass. Our study provides the first biochemical characterization of a multiprotein FA complex and suggests a connection between the BLM and FA pathways of genomic maintenance. The findings that FA proteins are part of a DNA-unwinding complex imply that FA proteins may participate in DNA repair.

X-linked inheritance of Fanconi anemia complementation group B

“…We immunopurified Fanconi anemia core complex from HeLa cells with an antibody to FANCA as described 3 . We identified FAAP95 by liquid chromatography and mass spectrometry analysis; the mass data are available on request.…”

“…The immunoprecipitation conditions were previously described 7 . We prepared nuclear and cytoplasmic extracts as described 3 . Owing to its low abundance, FAAP95 cannot be detected by direct immunoblotting analysis of the cell extract.…”

“…We transfected HeLa cells using siRNA oligonucleotides as described 3 . The FAAP95 siRNA oligonucleotide comprised a 19-nucleotide target sequence (5¢-UGUACUCAAGAGCCUUCAA-3¢) and was purchased from Dharmacon Research.…”

“…We showed that FAAP43 (also called FANCL) is a ubiquitin ligase and was defective in an individual affected with Fanconi anemia belonging to a new complementation group (FA-L) 3,7 (Fig. 1a).…”

The Fanconi Anemia/BRCA pathway: new faces in the crowd

“…BLM and topoisomerase III have also been found to associate with FA complex 1 (Meetei et al 2003b). BLM, topoisomerase III, and replication protein A associate with constituents of the FA complex 1 (the BRAFT complex) (Meetei et al 2003b).…”