V(D)J recombination and class switch recombination employ overlapping but distinct non-homologous end-joining (NHEJ) pathways to repair DNA double strand break (DSB) intermediates. 53BP1 is a DNA damage response protein that is rapidly recruited to sites of chromosomal DSBs, where it appears to function in a subset of ataxia-telangiectasia mutated (ATM) kinase, H2AX- and MDC1- dependent events1,2. A 53BP1 dependent end joining pathway has been described that is dispensable for V(D)J recombination but essential for class-switch recombination CSR3, 4. Here, we report a previously unrecognized defect in the joining phase of V(D)J recombination in 53BP1 deficient lymphocytes distinct from that found in classical NHEJ-, H2AX-, MDC1- and Atm-deficient mice. Absence of 53BP1 leads to impairment of distal V-DJ joining with extensive degradation of un-repaired coding ends and episomal signal joint reintegration at V(D)J junctions. This results in apoptosis, loss of T-cell receptor alpha locus integrity and lymphopenia. Further impairment of the apoptotic checkpoint causes propagation of lymphocytes bearing antigen receptor breaks. These data suggest a more general role for 53BP1 in maintaining genomic stability during long range joining of DNA breaks.

A-type lamins are intermediate filament proteins that provide a scaffold for protein complexes regulating nuclear structure and function. Mutations in the LMNA gene are linked to a variety of degenerative disorders termed laminopathies, whereas changes in the expression of lamins are associated with tumourigenesis. The molecular pathways affected by alterations of A-type lamins and how they contribute to disease are poorly understood. Here, we show that A-type lamins have a key role in the maintenance of telomere structure, length and function, and in the stabilization of 53BP1, a component of the DNA damage response (DDR) pathway. Loss of A-type lamins alters the nuclear distribution of telomeres and results in telomere shortening, defects in telomeric heterochromatin, and increased genomic instability. In addition, A-type lamins are necessary for the processing of dysfunctional telomeres by non-homologous end joining, putatively through stabilization of 53BP1. This study shows new functions for A-type lamins in the maintenance of genomic integrity, and suggests that alterations of telomere biology and defects in DDR contribute to the pathogenesis of lamin-related diseases.

DNA double strand breaks are generated by genotoxic agents and by cellular endonucleases as intermediates of several important physiologic processes. The cellular response to genotoxic DNA breaks includes the activation of transcriptional programs known primarily to regulate cell cycle checkpoints and cell survival1–5. DNA double strand breaks are generated in all developing lymphocytes during the assembly of antigen receptor genes, a process that is essential for normal lymphocyte development. Here we demonstrate that these physiologic DNA breaks activate a broad transcriptional program. This program transcends the canonical DNA double strand break response and includes many genes that regulate diverse cellular processes important for lymphocyte development. Moreover, the expression of several of these genes is regulated similarly in response to genotoxic DNA damage. Thus, physiologic DNA double strand breaks provide cues that can regulate cell-type-specific processes not directly involved in maintaining the integrity of the genome, and genotoxic DNA breaks could disrupt normal cellular functions by corrupting these processes.

Phosphorylation of DNA-PKcs is essential for activation of DNA damage repair and for the maintenance of tissue stem cell populations.

Previous work has shown that laulimalide, a sponge-derived natural product, resembles paclitaxel in enhancing tubulin assembly and in its effects on cellular microtubules. The two compounds, however, seem to have distinct binding sites on tubulin polymer. Nearly equimolar amounts of tubulin, laulimalide, and paclitaxel are recovered from microtubules formed with both drugs. In the present study, we searched for differences between laulimalide and paclitaxel in their interactions with tubulin polymer. Laulimalide was compared with paclitaxel and epothilone A, a natural product that competes with paclitaxel in binding to microtubules, for assembly properties at different temperatures and for effects of GTP and microtubule-associated proteins on assembly. Although minor differences were observed among the three drugs, their overall effects were highly similar, except that aberrant assembly products were observed more frequently with paclitaxel and that the polymers formed with laulimalide and epothilone A were more stable at 0°C. The most dramatic difference observed between laulimalide and epothilone A was that only laulimalide was able to enhance assembly synergistically with paclitaxel, as would be predicted if the two drugs bound at different sites in polymer. Because stoichiometric amounts of laulimalide and paclitaxel can cause extensive tubulin assembly, maximum synergy was observed at lower temperatures under reaction conditions in which each drug alone is relatively inactive. Laulimalide-induced assembly, like paclitaxel-induced assembly, was inhibited by drugs that inhibit tubulin assembly by binding at either the colchicine-or vinblastine-binding site. When radiolabeled GTP is present in a reaction mixture with either laulimalide or paclitaxel, nucleotide hydrolysis occurs with incorporation of radiolabeled GDP into polymer.