title: Initiation of V(D)J recombination in a cell-free system.
abstract: Cells performing V(D)J recombination make specific cuts in DNA at recombination signal sequences. Here, we show that nuclear extracts of pre-B cell lines carry out this specific cleavage. The products of cleavage are the same as found previously in thymocytes: full-length, blunt, 5'-phosphorylated signal ends, and covalently sealed (hairpin) coding ends. A complete signal sequence is required. Recombinant RAG1 protein greatly increases activity and complements an inactive extract from a RAG1 (-/-) pre-B cell line. When the extracts are fractionated, cleavage activity correlates with the presence of RAG2 protein. These results suggest that RAG1 and RAG2 are components of the V(D)J recombinase.
authors.affiliation: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: D C
authors.initials: DC
authors.lastname: van Gent
authors.forename: J F
authors.initials: JF
authors.lastname: McBlane
authors.forename: D A
authors.initials: DA
authors.lastname: Ramsden
authors.forename: M J
authors.initials: MJ
authors.lastname: Sadofsky
authors.forename: J E
authors.initials: JE
authors.lastname: Hesse
authors.forename: M
authors.initials: M
authors.lastname: Gellert
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology; D001483:Base Sequence; D002460:Cell Line; D002474:Cell-Free System; D004247:DNA / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004268:DNA-Binding Proteins; D005803:Genes, Immunoglobulin; D018398:Homeodomain Proteins; D008969:Molecular Sequence Data; D011506:Proteins / Q000235:genetics; D011948:Receptors, Antigen, T-Cell / Q000235:genetics*; D011995:Recombination, Genetic
chemical_list: D017931:DNA Primers; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D011506:Proteins; D011948:Receptors, Antigen, T-Cell; C064655:V(D)J recombination activating protein 2; C064658:RAG-1 protein; D004247:DNA