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The In Vivo Pattern of Binding of RAG1 and RAG2 to Antigen Receptor Loci

ABSTRACT: The critical initial step in V(D)J recombination, binding of RAG1 and RAG2 to recombination signal sequences flanking antigen receptor V, D, and J gene segments, has not previously been characterized in vivo. Here we demonstrate that RAG protein binding occurs in a highly focal manner to a small region of active chromatin encompassing Igκ and Tcrα J gene segments and Igh and Tcrβ J and J-proximal D gene segments. Formation of these small RAG-bound regions, which we refer to as recombination centers, occurs in a developmental stage- and lineage-specific manner. Each RAG protein is independently capable of specific binding within recombination centers. While RAG1 binding is restricted to regions containing recombination signal sequences, RAG2 binds extremely broadly in a pattern that mirrors that of trimethylated lysine 4 of histone 3. We propose that recombination centers coordinate V(D)J recombination by providing discrete sites within which gene segments are captured for recombination.

ORGANISM(S): Mus musculus

PROVIDER: GSE21207 | GEO | 2010/04/20

SECONDARY ACCESSION(S): PRJNA126295

REPOSITORIES: GEO

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The In Vivo Pattern of Binding of RAG1 and RAG2 to Antigen Receptor Loci

Project description:The critical initial step in V(D)J recombination, binding of RAG1 and RAG2 to recombination signal sequences flanking antigen receptor V, D, and J gene segments, has not previously been characterized in vivo. Here we demonstrate that RAG protein binding occurs in a highly focal manner to a small region of active chromatin encompassing IgÎº and TcrÎ± J gene segments and Igh and TcrÎ² J and J-proximal D gene segments. Formation of these small RAG-bound regions, which we refer to as recombination centers, occurs in a developmental stage- and lineage-specific manner. Each RAG protein is independently capable of specific binding within recombination centers. While RAG1 binding is restricted to regions containing recombination signal sequences, RAG2 binds extremely broadly in a pattern that mirrors that of trimethylated lysine 4 of histone 3. We propose that recombination centers coordinate V(D)J recombination by providing discrete sites within which gene segments are captured for recombination. RAG2 binding was analyzed in wild type, RAG2-/-Î², and D708A-RAG1-/-Î² thymocytes. Histone modification H3K4me3 was analyzed in D708A-RAG1-/-Î² and wild type thymocytes.

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RAG1 targeting in the genome is dominated by chromatin interactions mediated by the non-core regions of RAG1 and RAG2

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