title: VprBP binds full-length RAG1 and is required for B-cell development and V(D)J recombination fidelity.
abstract: The N-terminus of full-length RAG1, though dispensable for RAG1/2 cleavage activity, is required for efficient V(D)J recombination. This region supports RING E3 ubiquitin ligase activity in vitro, but whether full-length RAG1 functions as a single subunit or a multi-subunit E3 ligase in vivo is unclear. We show the multi-subunit cullin RING E3 ligase complex VprBP/DDB1/Cul4A/Roc1 associates with full-length RAG1 through VprBP. This complex is assembled into RAG protein-DNA complexes, and supports in-vitro ubiquitylation activity that is insensitive to RAG1 RING domain mutations. Conditional B lineage-specific VprBP disruption arrests B-cell development at the pro-B-to-pre-B cell transition, but this block is bypassed by expressing rearranged immunoglobulin transgenes. Mice with a conditional VprBP disruption show modest reduction of D-J(H) rearrangement, whereas V(H)-DJ(H) and V(κ)-J(κ) rearrangements are severely impaired. D-J(H) coding joints from VprBP-insufficent mice show longer junctional nucleotide insertions and a higher mutation frequency in D and J segments than normal. These data suggest full-length RAG1 recruits a cullin RING E3 ligase complex to ubiquitylate an unknown protein(s) to limit error-prone repair during V(D)J recombination.
authors.affiliation: Department of Medical Microbiology and Immunology, Creighton University Medical Center, Omaha, NE, USA.
authors.forename: Michele D
authors.initials: MD
authors.lastname: Kassmeier
authors.forename: Koushik
authors.initials: K
authors.lastname: Mondal
authors.forename: Victoria L
authors.initials: VL
authors.lastname: Palmer
authors.forename: Prafulla
authors.initials: P
authors.lastname: Raval
authors.forename: Sushil
authors.initials: S
authors.lastname: Kumar
authors.forename: Greg A
authors.initials: GA
authors.lastname: Perry
authors.forename: Dirk K
authors.initials: DK
authors.lastname: Anderson
authors.forename: Pawel
authors.initials: P
authors.lastname: Ciborowski
authors.forename: Sarah
authors.initials: S
authors.lastname: Jackson
authors.forename: Yue
authors.initials: Y
authors.lastname: Xiong
authors.forename: Patrick C
authors.initials: PC
authors.lastname: Swanson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology*; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases; D019076:Transgenes; D060152:V(D)J Recombination
chemical_list: D002352:Carrier Proteins; D018398:Homeodomain Proteins; C064658:RAG-1 protein; D017346:Protein Serine-Threonine Kinases; C571226:VprBP protein, mouse