GSE21207
The In Vivo Pattern of Binding of RAG1 and RAG2 to Antigen Receptor Loci
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 Ig? and Tcra J gene segments and Igh and Tcrß 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. Overall design: RAG2 binding was analyzed in wild type, RAG2-/-ß, and D708A-RAG1-/-ß thymocytes. Histone modification H3K4me3 was analyzed in D708A-RAG1-/-ß and wild type thymocytes.