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The Conserved ATM Kinase RAG2-S365 Phosphorylation Site Limits Cleavage Events in Individual Cells Independent of Any

Susannah L Hewitt1, Jason B Wong1, Ji-Hoon Lee2

  • 1Department of Pathology, New York University School of Medicine, New York, NY 10016, USA.

Cell Reports
|October 26, 2017
PubMed

Insights

The RAG1/2 recombinase

Area of Science:

  • Molecular Biology
  • Genetics
  • Immunology

Background:

  • Off-target DNA cleavage by the RAG1/2 recombinase is linked to lymphoid malignancies.
  • Understanding RAG cleavage deregulation is hindered by confounding DNA repair defects.

Purpose of the Study:

  • To investigate the role of RAG2 phosphorylation in controlling RAG-mediated DNA cleavage.
  • To establish deregulated cleavage as a driver of chromosomal instability independent of repair defects.

Main Methods:

  • Site-directed mutagenesis of RAG2 phosphorylation site (S365A).
  • Analysis of RAG-mediated DNA breaks and chromosomal translocations.
  • Assessment of ATM kinase inactivation and RAG2 phosphomimetic mutations (S365E).

Main Results:

  • Mutation of RAG2 S365 to alanine causes bi-allelic, bi-locus breaks and reciprocal translocations.
  • Deregulated RAG cleavage, not repair defects, drives chromosomal instability.
  • RAG2-S365E phosphomimetic mutation rescues ATM-inactivation-induced translocations.

Conclusions:

  • ATM-mediated phosphorylation of RAG2 is crucial for feedback control of RAG cleavage.
  • This phosphorylation mechanism maintains genome stability by preventing deregulated cleavage.
  • RAG2 phosphorylation by ATM is a key regulatory pathway in V(D)J recombination and genome integrity.
Keywords:
ATMRAG cleavage regulationRAG2S365V(D)J recombinationdeveloping lymphocytesgenome stabilityreciprocal translocations

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