DNA ligase IV mutations confer shorter lifespan and increased sensitivity to nutrient stress in Drosophila melanogaster

The nonhomologous end-joining pathway is a primary DNA double-strand break repair pathway in eukaryotes. DNA ligase IV (Lig4) catalyzes the final step of DNA end ligation in this pathway. Partial loss of Lig4 in mammals causes Lig4 syndrome, while complete loss is embryonically lethal. DNA ligase 4 (DNAlig4) null Drosophila melanogaster is viable, but sensitive to ionizing radiation during early development. We proposed to explore if DNAlig4 loss induced other long-term sensitivities and defects in D. melanogaster. We demonstrated that DNAlig4 mutant strains had decreased lifespan and lower resistance to nutrient deprivation, indicating Lig4 is required for maintaining health and longevity in D. melanogaster.

A XRCC4 mutant mouse, a model for human X4 syndrome, reveals interplays with Xlf, PAXX, and ATM in lymphoid development

We developed a Xrcc4M61R separation of function mouse line to overcome the embryonic lethality of Xrcc4 deficient mice. XRCC4M61R protein does not interact with Xlf, thus obliterating XRCC4-Xlf filament formation while preserving the ability to stabilize DNA Ligase IV. X4M61R mice, which are DNA repair deficient, phenocopy the Nhej1-/- (known as Xlf -/-) setting with a minor impact on the development of the adaptive immune system. The core NHEJ DNA repair factor XRCC4 is therefore not mandatory for V(D)J recombination aside from its role in stabilizing DNA ligase IV. In contrast, Xrcc4M61R mice crossed on Paxx-/-, Nhej1-/-, or Atm-/- backgrounds are severely immunocompromised, owing to aborted V(D)J recombination as in Xlf-Paxx and Xlf-Atm double KO settings. Furthermore, massive apoptosis of post-mitotic neurons causes embryonic lethality of Xrcc4M61R -Nhej1-/- double mutants. These in vivo results reveal new functional interplays between XRCC4 and PAXX, ATM and Xlf in mouse development and provide new insights in the understanding of the clinical manifestations of human XRCC4 deficient condition, in particular its absence of immune deficiency.

Haploidentical Transplantation in a DNA Ligase IV Deficiency Patient: A Case Report and Review of The Literature

Background: DNA ligase IV (LIG4) deficiency is a rare autosomal recessive disorder caused by mutations in the DNA LIG4 gene. Nowadays hematopoietic stem cell transplantation(HSCT) is the most effective treatment option. Matched sibling or unrelated donor was the best choice for those patients. However, it will be a problem for LIG4 deficiency patients without proper donor as above but needed urgent transplantation because of infection or bone marrow failure. Furthermore, mixed donor chimerism after transplantation is more common in LIG4 deficiency patients because of reduced intensity conditioning(RIC) regimen. How to deal with those problems? Here we report a case which the patient received haploidentical HSCT and got complete donor chimerism after donor stem cell infusion. Results:The patient was diagnosed as LIG4 deficiency at 8-month-old. At 14 months of age, she received a T cell receptor(TCR)α/β and CD19+ B cells depleted graft from his haploidentical father, followed a RIC regimen with no additional graft versus host disease(GvHD) prophylaxis. Engraftment was as usual. However, mixed donor chimerism occurred after transplantation and viremia persisted. Cryopreserved donor cell infusion was initiated. The chimerism grew up steadily and viremia disappeared at four months post transplantation. Conclusions: This case report gives an example of successful haploidentical transplantation and donor stem cell infusion as a treatment option in a mixed donor chimerism situation after HSCT in LIG4 deficiency patients.

NAD+ is not utilized as a co-factor for DNA ligation by human DNA ligase IV

As nucleotidyl transferases, formation of a covalent enzyme-adenylate intermediate is a common first step of all DNA ligases. While it has been shown that eukaryotic DNA ligases utilize ATP as the adenylation donor, it was recently reported that human DNA ligase IV can also utilize NAD+ and, to a lesser extent ADP-ribose, as the source of the adenylate group and that NAD+, unlike ATP, enhances ligation by supporting multiple catalytic cycles. Since this unexpected finding has significant implications for our understanding of the mechanisms and regulation of DNA double strand break repair, we attempted to confirm that NAD+ and ADP-ribose can be used as co-factors by human DNA ligase IV. Here, we provide evidence that NAD+ does not enhance ligation by pre-adenylated DNA ligase IV, indicating that this co-factor is not utilized for re-adenylation and subsequent cycles of ligation. Moreover, we find that ligation by de-adenylated DNA ligase IV is dependent upon ATP not NAD+ or ADP-ribose. Thus, we conclude that human DNA ligase IV cannot use either NAD+ or ADP-ribose as adenylation donor for ligation.