title: Domain-Specific and Stage-Intrinsic Changes in Tcrb Conformation during Thymocyte Development.
abstract: Considerable cross-talk exists between mechanisms controlling genome architecture and gene expression. AgR loci are excellent models for these processes because they are regulated at both conformational and transcriptional levels to facilitate their assembly by V(D)J recombination. Upon commitment to the double-negative stage of T cell development, Tcrb adopts a compact conformation that promotes long-range recombination between Vβ gene segments (Trbvs) and their DβJβ targets. Formation of a functional VβDβJβ join signals for robust proliferation of double-negative thymocytes and their differentiation into double-positive (DP) cells, where Trbv recombination is squelched (allelic exclusion). DP differentiation also is accompanied by decontraction of Tcrb, which has been thought to separate the entire Trbv cluster from DβJβ segments (spatial segregation-based model for allelic exclusion). However, DP cells also repress transcription of unrearranged Trbvs, which may contribute to allelic exclusion. We performed a more detailed study of developmental changes in Tcrb topology and found that only the most distal portion of the Trbv cluster separates from DβJβ segments in DP thymocytes, leaving most Trbvs spatially available for rearrangement. Preferential dissociation of distal Trbvs is independent of robust proliferation or changes in transcription, chromatin, or architectural factors, which are coordinately regulated across the entire Trbv cluster. Segregation of distal Trbvs also occurs on alleles harboring a functional VβDβJβ join, suggesting that this process is independent of rearrangement status and is DP intrinsic. Our finding that most Trbvs remain associated with DβJβ targets in DP cells revises allelic exclusion models from their current conformation-dominant to a transcription-dominant formulation.
authors.affiliation: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110;
authors.forename: Kinjal
authors.initials: K
authors.lastname: Majumder
authors.affiliation: Division of Cancer Pathobiology, Department of Pathology and Laboratory Medicine, Center for Childhood Cancer Research, Children's Hospital of Philadelphia, Philadelphia, PA 19104; and Abramson Family Cancer Research Institute, Cell and Molecular Biology Graduate Program, Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Levi J
authors.initials: LJ
authors.lastname: Rupp
authors.affiliation: Division of Cancer Pathobiology, Department of Pathology and Laboratory Medicine, Center for Childhood Cancer Research, Children's Hospital of Philadelphia, Philadelphia, PA 19104; and Abramson Family Cancer Research Institute, Cell and Molecular Biology Graduate Program, Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Katherine S
authors.initials: KS
authors.lastname: Yang-Iott
authors.affiliation: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110;
authors.forename: Olivia I
authors.initials: OI
authors.lastname: Koues
authors.affiliation: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110;
authors.forename: Katherine E
authors.initials: KE
authors.lastname: Kyle
authors.affiliation: Division of Cancer Pathobiology, Department of Pathology and Laboratory Medicine, Center for Childhood Cancer Research, Children's Hospital of Philadelphia, Philadelphia, PA 19104; and Abramson Family Cancer Research Institute, Cell and Molecular Biology Graduate Program, Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104 eoltz@wustl.edu bassing@mail.med.upenn.edu.
authors.forename: Craig H
authors.initials: CH
authors.lastname: Bassing
authors.affiliation: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110; eoltz@wustl.edu bassing@mail.med.upenn.edu.
authors.forename: Eugene M
authors.initials: EM
authors.lastname: Oltz
mesh_terms: D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation / Q000276:immunology; D049109:Cell Proliferation / Q000235:genetics; D056743:Cyclin D3 / Q000235:genetics; D015333:Gene Rearrangement, beta-Chain T-Cell Antigen Receptor / Q000235:genetics*; D019674:Genes, T-Cell Receptor beta / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D018398:Homeodomain Proteins / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics*; D017422:Sequence Analysis, DNA; D060168:Thymocytes / Q000166:cytology*; D060152:V(D)J Recombination / Q000235:genetics*
chemical_list: C534697:Ccnd3 protein, mouse; D056743:Cyclin D3; D018398:Homeodomain Proteins; D016693:Receptors, Antigen, T-Cell, alpha-beta; C064658:RAG-1 protein