Binding and allosteric transmission of histone H3 Lys-4 trimethylation to the recombinase RAG-1 are separable functions of the RAG-2 plant homeodomain finger

Abstract

<jats:p>V(D)J recombination is initiated by the recombination-activating gene protein (RAG) recombinase, consisting of RAG-1 and RAG-2 subunits. The susceptibility of gene segments to cleavage by RAG is associated with gene transcription and with epigenetic marks characteristic of active chromatin, including histone H3 trimethylated at lysine 4 (H3K4me3). Binding of H3K4me3 by a plant homeodomain (PHD) in RAG-2 induces conformational changes in RAG-1, allosterically stimulating substrate binding and catalysis. To better understand the path of allostery from the RAG-2 PHD finger to RAG-1, here we employed phylogenetic substitution. We observed that a chimeric RAG-2 protein in which the mouse PHD finger is replaced by the corresponding domain from the shark <jats:italic>Chiloscyllium punctatum</jats:italic> binds H3K4me3 but fails to transmit an allosteric signal, indicating that binding of H3K4me3 by RAG-2 is insufficient to support recombination. By substituting residues in the <jats:italic>C. punctatum</jats:italic> PHD with the corresponding residues in the mouse PHD and testing for rescue of allostery, we demonstrate that H3K4me3 binding and transmission of an allosteric signal to RAG-1 are separable functions of the RAG-2 PHD finger.</jats:p>

The Dm element located in the Igκ locus is hypermethylated in primary RAG- deficient pre-B cells

“…The non-core region (384-527 residues) of RAG2 contains a plant homeodomain nger (PHD) that speci cally recognizes histone H3 trimethylated at lysine 4 (H3K4me3). This helps enhance the catalytic activity of the RAG proteins and guides RAG2 to active chromatin regions [9,10]. In addition, 350-383 residues of RAG2 also restricts RAG recombinase activity and is responsible for the Igκ locus demethylation in pre-B cells [11].However, the methylation status of Dm element remains unknown, and it is unclear whether RAG proteins are involved in its methylation changes in pre-B cells .…”