A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo

Abstract

The molecular complexes involved in the nonhomologous end-joining process that resolves recombination-activating gene (RAG)-induced double-strand breaks and results in V(D)J gene rearrangements vary during mammalian ontogeny. In the mouse, the first immunoglobulin gene rearrangements emerge during midgestation periods, but their repertoires have not been analyzed in detail. We decided to study the postgastrulation DJ_H joints and compare them with those present in later life. The embryo DJ_H joints differed from those observed in perinatal life by the presence of short stretches of nontemplated (N) nucleotides. Whereas most adult N nucleotides are introduced by terminal deoxynucleotidyl transferase (TdT), the embryo N nucleotides were due to the activity of the homologous DNA polymerase μ (Polμ), which was widely expressed in the early ontogeny, as shown by analysis of Polμ^−/− embryos. Based on its DNA-dependent polymerization ability, which TdT lacks, Polμ also filled in small sequence gaps at the coding ends and contributed to the ligation of highly processed ends, frequently found in the embryo, by pairing to internal microhomology sites. These findings show that Polμ participates in the repair of early-embryo, RAG-induced double-strand breaks and subsequently may contribute to preserve the genomic stability and cellular homeostasis of lymphohematopoietic precursors during development.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/ .

ACKNOWLEDGMENTS

This work was supported by grants SAF2007-65265, BFU2006-14390/BMC, and CSD2007-00015 from the Ministry of Education of Spain; by the Fundación Mutua Madrileña; and by S-SAL-0304-2006 from the Community of Madrid. The Centro de Biología Molecular Severo Ochoa receives institutional funding from Fundación Ramón Areces. B.G.-L. and N.S. are recipients of fellowships from Instituto de Salud Carlos III, and G.T. has a fellowship from the Spanish Ministry of Science and Technology.

We thank Fernando Martínez and Mario Alía for expert technical assistance, David Abia for statistical support, and Susana Morales and Daniel Lucas for contributions in the early stages of the work.