Aberrant V(D)J Recombination: Misrepair of DNA Breaks and Implications For Lymphomagenesis.

Abstract

Canonical non-homologous end joining (C-NHEJ), a major DNA double-strand break (DSB) repair pathway, is required for the repair of general DSBs as well as programmed breaks generated during antigen receptor gene rearrangements by V(D)J recombination. During this process, the lymphocyte-specific RAG1/2 endonuclease generates DSBs at numerous V, D and J genomic segments. The DNA ends are maintained in a postcleavage complex to facilitate end joining by C-NHEJ proteins, including Artemis, a DNA nuclease. Proper end joining is essential for lymphocyte development, immune diversification and to suppress structural chromosomal aberrations. Patients with null and hypomorphic mutations in RAG, Artemis and other C-NHEJ genes develop immunodeficiency disorders. Hypomorphic Artemis mutations that result in loss of the C-terminus are also associated with lymphomas harboring chromosomal translocations at the antigen receptor loci. The mechanisms that promote genomic instability in these patients are not entirely clear. This dissertation is focused on elucidating the roles of RAG1 and Artemis in preventing genomic instability during V(D)J recombination. These studies provide the first in vivo evidence that hypomorphic Rag1 and Artemis mutations result in unique molecular defects characterized by chromosomal translocations at the RAG-induced breaks in non-malignant cells. My findings suggest that RAG1 and Artemis play important roles in maintaining postcleavage complex stability. Disruption of these complexes in the hypomorphic mutant lymphocytes results in premature release of DNA ends that may engage in aberrant V(D)J recombination. Recurrent chromosomal translocations are also a hallmark of sporadic human hematological malignancies, and of progenitor-B cell lymphomas in mice with combined C-NHEJ and tumor suppressor p53 deficiencies. Distinct alternative end joining (A-EJ) mechanisms have been implicated in mediating these translocations. The mechanisms that activate A-EJ are poorly characterized. My studies have demonstrated that induction of DNA damage response triggers alternative end-joining (A-EJ) in lymphocytes in the absence of Artemis resulting in chromosomal translocations at RAG-induced breaks. In summary, I have identified novel functions for RAG and Artemis in maintaining chromosomal integrity during V(D)J recombination. These studies have improved our understanding of the origin of potentially oncogenic chromosomal translocations and have important clinical implications for both inherited cancer predisposition and sporadic tumors.