| First Author | Zhu Y | Year | 2023 |
| Journal | Proc Natl Acad Sci U S A | Volume | 120 |
| Issue | 25 | Pages | e2221894120 |
| PubMed ID | 37307443 | Mgi Jnum | J:360028 |
| Mgi Id | MGI:7567341 | Doi | 10.1073/pnas.2221894120 |
| Citation | Zhu Y, et al. (2023) Phosphorylation of DNA-PKcs at the S2056 cluster ensures efficient and productive lymphocyte development in XLF-deficient mice. Proc Natl Acad Sci U S A 120(25):e2221894120 |
| abstractText | The nonhomologous end-joining (NHEJ) pathway is a major DNA double-strand break repair pathway in mammals and is essential for lymphocyte development. Ku70 and Ku80 heterodimer (KU) initiates NHEJ, thereby recruiting and activating the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs). While DNA-PKcs deletion only moderately impairs end-ligation, the expression of kinase-dead DNA-PKcs completely abrogates NHEJ. Active DNA-PK phosphorylates DNA-PKcs at two clusters-PQR around S2056 (S2053 in mouse) and ABCDE around T2609. Alanine substitution at the S2056 cluster moderately compromises end-ligation on plasmid-based assays. But, mice carrying alanine substitution at all five serine residues within the S2056 cluster (DNA-PKcs(PQR/PQR)) display no defect in lymphocyte development, leaving the physiological significance of S2056 cluster phosphorylation elusive. Xlf is a nonessential NHEJ factor. Xlf (-/-) mice have substantial peripheral lymphocytes that are completely abolished by the loss of DNA-PKcs, the related ATM kinases, other chromatin-associated DNA damage response factors (e.g., 53BP1, MDC1, H2AX, and MRI), or RAG2-C-terminal regions, suggesting functional redundancy. While ATM inhibition does not further compromise end-ligation, here we show that in XLF-deficient background, DNA-PKcs S2056 cluster phosphorylation is critical for normal lymphocyte development. Chromosomal V(D)J recombination from DNA-PKcs(PQR/PQR)Xlf (-/-) B cells is efficient but often has large deletions that jeopardize lymphocyte development. Class-switch recombination junctions from DNA-PKcs(PQR/PQR)Xlf (-/-) mice are less efficient and the residual junctions display decreased fidelity and increased deletion. These findings establish a role for DNA-PKcs S2056 cluster phosphorylation in physiological chromosomal NHEJ, implying that S2056 cluster phosphorylation contributes to the synergy between XLF and DNA-PKcs in end-ligation. |
