Disease relevance of NHEJ1

• Here, we describe a patient with T(-)B(-) severe combined immunodeficiency, whose cells have defects closely resembling those of NHEJ-defective rodent cells [1].
• Although nonhomologous end-joining (NHEJ) deficiency has been shown to accelerate lymphoma formation in mice, its role in suppressing tumors in cells that do not undergo V(D)J recombination is unclear [2].
• Together, these findings support the view that loss of a single lig4 allele results in NHEJ activity being sufficiently reduced to engender chromosomal aberrations that drive non-lymphoid tumorigenesis [2].
• In the non-homologous end-joining (NHEJ) pathway, a polymorphism in LIG4 (T>C at nt 1977) was associated with a decrease in breast cancer risk [P=0.09; OR CC versus TT=0.7 (0.4-1.0)] [3].
• METHODS AND MATERIALS: The expression of homologous recombination (HR) and non-homologous recombination (NHEJ) genes following gas hypoxia (0.2%) or exposure to HIF1alpha-inducing agent, CoCl2 (100 microM), was determined for normal diploid fibroblasts (GM05757) and the pre-malignant and malignant prostate cell lines, BPH-1, 22RV-1, DU145 and PC3 [4].

High impact information on NHEJ1

• Central to NHEJ is the protein complex containing DNA Ligase IV and XRCC4 [5].
• Cernunnos/XLF represents a novel DNA repair factor essential for the NHEJ pathway [6].
• Cernunnos, a novel nonhomologous end-joining factor, is mutated in human immunodeficiency with microcephaly [6].
• We show that XLF directly interacts with the XRCC4-Ligase IV complex in vitro and in vivo and that siRNA-mediated downregulation of XLF in human cell lines leads to radiosensitivity and impaired NHEJ [5].
• The t-loop structure of mammalian telomeres is thought to repress nonhomologous end joining (NHEJ) at natural chromosome ends [7].

Chemical compound and disease context of NHEJ1

• Based on comprehensive NHEJ gene profiles, this study provides new insights to suggest the role of the NHEJ pathway in breast cancer development and supports the possibility that breast cancer is initiated by estrogen exposure, which causes DNA DSBs [8].

Biological context of NHEJ1

• Indeed, we show that a member of this family encoded by a previously uncharacterized open-reading frame in the Schizosaccharomyces pombe genome is required for NHEJ in this organism [9].
• We speculate that Cernunnos-XLF may have contributed to the increased number of brain cells in humans by efficient double strand break repair, which helps to prevent frequent apoptosis of neuronal progenitors and aids mitotic cell cycle progression [10].
• The involvement of inositol phosphate in DNA-PK-dependent NHEJ is of particular interest since the catalytic domain of DNA-PKcs is similar to that found in the phosphatidylinositol 3 (PI 3)-kinase family [11].
• Chromosomal double-strand breaks (DSBs) in mammalian cells are repaired by either homology-directed repair (HDR), using a homologous sequence as a repair template, or nonhomologous end-joining (NHEJ), which often involves sequence alterations at the DSB site [12].
• Neither sister-chromatid exchange nor gene-targeting frequencies show a dependence on these NHEJ proteins [12].

Anatomical context of NHEJ1

• NHEJ also functions in developing lymphocytes to repair strand breaks that occur during V(D)J recombination, the site-specific recombination process that provides for the assembly of functional antigen-receptor genes [13].
• Non-homologous end joining (NHEJ) is one of two pathways responsible for the repair of double-strand breaks in eukaryotic cells [14].
• Using human whole cell extracts prepared by the method of Baumann and West (1998), we have described a cell-free system for NHEJ that joins both compatible and noncompatible DNA ends (Budman and Chu, 2005) [15].
• All mammalian mutants in the NHEJ pathway demonstrate a lack of B and T lymphocytes and ionizing radiation sensitivity [16].
• In addition, we show that potential anti-cancer drugs (LY-294002 and vanillin) that inhibit the family of phosphatidylinositol 3 kinases that include the NHEJ protein, DNA-PKCS act in synergy with TSA to reduce the viability of HeLa cells in tissue culture presenting the possibility of using the two drugs in combination to treat cancer [17].

Associations of NHEJ1 with chemical compounds

• We show that polynucleotide kinase (PNK), with its ability to process ionizing radiation-induced 5'-OH and 3'-phosphate DNA termini, functions in NHEJ via an FHA-dependent interaction with CK2-phosphorylated Xrcc4 [18].
• By designing two truncated forms of pol lambda, we also show that the unique proline-rich region in pol lambda plays a role in limiting strand displacement synthesis, a feature that may help its participation in in vivo NHEJ [14].
• By using a mutational approach, we have identified a consensus Dun1p phosphorylation site in Nej1p, and mutation of conserved serine residues within it leads to decreased NHEJ efficiency [19].
• Both HR and NHEJ protected cells from the lethal effects of hydroxyurea, and this agent also increased the frequency of recombination mediated by both homologous and nonhomologous exchanges [20].
• Plasma XLFP were identified by sodium dodecyl sulfate (SDS)-agarose electrophoresis and Western blotting and quantitated by gel scanning [21].

Physical interactions of NHEJ1

• We show here that Cernunnos physically interacts with the XRCC4 x DNA-ligase IV complex [22].

Enzymatic interactions of NHEJ1

• Here we demonstrate that purified pol lambda can efficiently catalyze gap-filling synthesis on DNA substrates mimicking NHEJ [14].

Regulatory relationships of NHEJ1

• Based on these results, we hypothesize that Ad-mda7 sensitizes NSCLC cells to ionizing radiation by suppressing the activity of NHEJ, a pathway essential for repair of radiation-induced DSBs [23].
• These results support a model in which DNA-PKcs plays a central role in regulating the processing of ends for NHEJ [24].
• Junctions were repaired in the same manner in MO59K extracts in which accurate NHEJ was inactivated by inhibition of Ku70 or DNA-PK(cs) [25].

Other interactions of NHEJ1

• We find that all hybrids in vivo require DNA ligase IV in human cells, which is the final component of the NHEJ pathway [26].
• These results suggest that ATM activity is required for the residual transduction observed in the NHEJ-deficient cells [27].
• In addition, the NHEJ pathway per se was marginally affected by BLM deficiency, as evidenced by x-ray sensitivity and I-SceI-based DSB repair assays [28].
• The rates of appearance and dissolution of NHEJ proteins paralleled that of histone variant H2AX phosphorylation and dephosphorylation [29].
• The discovery of homologues from the yeast Saccharomyces cerevisiae of the human Ku DNA-end-binding proteins (HDF1 and KU80) has established that this organism is capable of non-homologous double-strand end joining (NHEJ), a form of DNA double-strand break repair (DSBR) active in mammalian V(D)J recombination [30].

Analytical, diagnostic and therapeutic context of NHEJ1

• Furthermore, our data reveal that XLF family proteins can bind to DNA and directly interact with the ligase IV-XRCC4 complex to promote DSB ligation [9].
• Moreover, a combination of sensitive methods of sequence analysis revealed that Cernunnos can be associated with the XRCC4 family of proteins and that it corresponds to the genuine homolog of the yeast Nej1 protein [22].
• The expression level and migration of key proteins from the nonhomologous end-joining (NHEJ) pathway was evaluated by Western blot analysis on cases and controls [31].
• Chromatin immunoprecipitation showed that Sem1 is recruited along with the 19S and 20S proteasomes to a DSB in vivo, and this recruitment is dependent on components of both the HR and NHEJ repair pathways, suggesting a direct role of the proteasome in DSB repair [32].
• Using immunofluorescence detection of gamma-H2AX nuclear foci as a novel approach for monitoring the repair of DSBs, we show here that NHEJ-defective hamster cells (CHO mutant V3 cells) have strongly reduced repair in all cell cycle phases after 1 Gy of irradiation [33].

References