title: Characterization of BTBD1 and BTBD2, two similar BTB-domain-containing Kelch-like proteins that interact with Topoisomerase I.
abstract: BACKGROUND
  Two-hybrid screening for proteins that interact with the core domain of human topoisomerase I identified two novel proteins, BTBD1 and BTBD2, which share 80% amino acid identities.
  
  
  RESULTS
  The interactions were confirmed by co-precipitation assays demonstrating the physical interaction of BTBD1 and BTBD2 with 100 kDa topoisomerase I from HeLa cells. Deletion mapping using two-hybrid and GST-pulldown assays demonstrated that less than the C-terminal half of BTBD1 is sufficient for binding topoisomerase I. The topoisomerase I sequences sufficient to bind BTBD2 were mapped to residues 215 to 329. BTBD2 with an epitope tag localized to cytoplasmic bodies. Using truncated versions that direct BTBD2 and TOP1 to the same cellular compartment, either the nucleus or the cytoplasm, co-localization was demonstrated in co-transfected Hela cells. The supercoil relaxation and DNA cleavage activities of topoisomerase I in vitro were affected little or none by co-incubation with BTBD2. Northern analysis revealed only a single sized mRNA for each BTBD1 and BTBD2 in all human tissues tested. Characterization of BTBD2 mRNA revealed a 255 nucleotide 90% GC-rich region predicted to encode the N-terminus. BTBD1 and BTBD2 are widely if not ubiquitously expressed in human tissues, and have two paralogs as well as putative orthologs in C. elegans and D. melanogaster.
  
  
  CONCLUSIONS
  BTBD1 and BTBD2 belong to a small family of uncharacterized proteins that appear to be specific to animals. Epitope-tagged BTBD2 localized to cytoplasmic bodies. The characterization of BTBD1 and BTBD2 and their interaction with TOP1 is underway.
authors.affiliation: Department of Biochemistry and Molecular Biology Uniformed Services University of the Health Sciences 4301 Jones Bridge Road, Bethesda, MD 20814-4799, USA. lixu@usuhs.mil
authors.forename: Lixin
authors.initials: L
authors.lastname: Xu
authors.forename: Lihong
authors.initials: L
authors.lastname: Yang
authors.forename: Keiko
authors.initials: K
authors.lastname: Hashimoto
authors.forename: Melvin
authors.initials: M
authors.lastname: Anderson
authors.forename: Glenda
authors.initials: G
authors.lastname: Kohlhagen
authors.forename: Yves
authors.initials: Y
authors.lastname: Pommier
authors.forename: Peter
authors.initials: P
authors.lastname: D'Arpa