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

J Biol Chem. 2020 Jul 3;295(27):9052-9060. doi: 10.1074/jbc.RA120.014382. Epub 2020 May 15.

Abstract

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 Chiloscyllium punctatum 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 C. punctatum 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.

Keywords: DNA recombination; H3K4me3; PHD finger; V(D)J rearrangement; allosteric regulation; conformational change; epigenetics; histone methylation; recombination-activating (RAG) recombinase.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Allosteric Regulation / genetics
  • Allosteric Regulation / physiology
  • Animals
  • Binding Sites
  • Chromatin / metabolism
  • DNA-Binding Proteins / metabolism*
  • Histones / metabolism*
  • Histones / physiology
  • Homeodomain Proteins / metabolism*
  • Lysine / metabolism
  • Methylation
  • Mice
  • Phylogeny
  • Protein Binding
  • Recombinases / metabolism
  • Sharks / metabolism
  • Substrate Specificity
  • V(D)J Recombination / genetics
  • V(D)J Recombination / physiology
  • VDJ Recombinases / metabolism

Substances

  • Chromatin
  • DNA-Binding Proteins
  • Histones
  • Homeodomain Proteins
  • Recombinases
  • V(D)J recombination activating protein 2
  • histone H3 trimethyl Lys4
  • RAG-1 protein
  • VDJ Recombinases
  • Lysine

Associated data

  • PDB/2V89