Hypomorphic Lig4 Impacts on Igh Class Switching and Translocations.

Abstract 3373

The DNA Ligase IV (Lig4) protein is an essential component of the non-homologous end-joining (NHEJ) repair pathway that plays a key role in the repair of general DNA double strand breaks (DSBs) and those generated during V(D)J recombination and IgH Class Switch Recombination (CSR) in B lymphocytes. Hypomorphic mutations in the Lig4 gene in humans underlies the DNA Lig4 Syndrome, which is associated with radiosensitivity, developmental delay, growth defects, facial dysmorphism and variable degrees of immunodeficiencies. We recently described a mouse model carrying a specific Lig4 hypomorphic mutation in both copies of the Lig4 coding exon that generates an arginine to histidine amino acid #278 (R278H) replacement that corresponds to the first Lig4 mutation reported in humans. Our studies revealed that similar to human patients, the phenotype of homozygous mutant mice Lig4R278H/R278H (Lig4R/R) includes growth retardation, significant decreased life span, severe cellular sensitivity to ionizing radiation, and a severe but incomplete block in T and B cell development. Peripheral T lymphocytes show an activated and anergic phenotype, reduced viability, and a restricted repertoire, reminiscent of human leaky SCID, and high levels of genomic instability associated with increased incidence of thymic and peripheral T lymphomas. Severe defects in B lymphocyte developmental functions include repertoire restriction during aging, spontaneous low affinity antibody production and inability mount high affinity antibody responses. Our studies now reveal that introduction of IgH and IgL knock-in (HL) variable region exon into the R/R mice leads to significant increase in mature B lymphocytes in peripheral lymphoid organs, directly demonstrating that the overall severe reduction in B lymphocytes in the R/R mice is in part due to a block in their developmental progression. We further observe a mild reduction in IgH class switching in the in vitro CSR activated R/RHL B lymphocytes, accompanied by an unusual increase in IgH specific DSBs and translocations, at levels significantly higher than what we previously observed in Lig4 or XRCC4 deficient peripheral B cells, and lower than expected general DSBs. The implications of our findings in the context of B lymphocyte development and CSR, potential altered utilization of alternative end-joining DSB repair mechanisms and on B lymphomagenesis will be discussed.