TY - JOUR
T1 - Protein phosphatase 1 and phosphatase 1 nuclear targeting subunit-dependent regulation of DNA-dependent protein kinase and non-homologous end joining
AU - Zhu, Songli
AU - Fisher, Laura A.
AU - Bessho, Tadayoshi
AU - Peng, Aimin
N1 - Funding Information:
We thank Dr Jay Reddy (University of Nebraska-Lincoln) for technical support with X-ray irradiation; Dr Li Lan (University of Pittsburgh) for assistance with imaging; Drs Jeremy Stark (City of Hope) and Maria Jasin (Memorial Sloan Kettering Cancer Center) for reagents; Drs Gregory Oakley and Ali Nawshad for stimulating discussions; Dr James Wahl and Nicholas Eurek for assisting with the microscopic studies. The UNMC Advanced Microscopy Core Facility was supported by the Nebraska Research Initiative, the Fred and Pamela Buffett Cancer Center Support Grant (P30CA036727), and an Institutional Development Award (IDeA) from the NIGMS of the NIH (P30GM106397).
Funding Information:
National Institutes of Health [1R01CA172574 to A.P.]. Funding for open access charge: National Institutes of Health [1R01CA172574 to A.P.]. Conflict of interest statement. None declared.
Publisher Copyright:
© The Author(s) 2017.
PY - 2017/10/1
Y1 - 2017/10/1
N2 - DNA-dependent protein kinase catalytic subunit (DNA-PKcs) plays a key role in mediating nonhomologous end joining (NHEJ), a major repair pathway for DNA double-strand breaks (DSBs). The activation, function and dynamics of DNA-PKcs is regulated largely by its reversible phosphorylation at numerous residues, many of which are targeted by DNA-PKcs itself. Interestingly, these DNA-PKcs phosphorylation sites function in a distinct, and sometimes opposing manner, suggesting that they are differentially regulated via complex actions of both kinases and phosphatases. In this study we identified several phosphatase subunits as potential DSB-associated proteins. In particular, protein phosphatase 1 (PP1) is recruited to a DSB-mimicking substrate in Xenopusegg extracts and sites of laser microirradiation in human cells. Depletion of PP1 impairs NHEJ in both Xenopus egg extracts and human cells. PP1 binds multiple motifs of DNA-PKcs, regulates DNA-PKcs phosphorylation, and is required for DNA-PKcs activation after DNA damage. Interestingly, phosphatase 1 nuclear targeting subunit (PNUTS), an inhibitory regulator of PP1, is also recruited to DNA damage sites to promote NHEJ. PNUTS associates with the DNA-PK complex and is required for DNA-PKcs phosphorylation at Ser-2056 and Thr-2609. Thus, PNUTS and PP1 together finetune the dynamic phosphorylation of DNA-PKcs after DNA damage to mediate NHEJ.
AB - DNA-dependent protein kinase catalytic subunit (DNA-PKcs) plays a key role in mediating nonhomologous end joining (NHEJ), a major repair pathway for DNA double-strand breaks (DSBs). The activation, function and dynamics of DNA-PKcs is regulated largely by its reversible phosphorylation at numerous residues, many of which are targeted by DNA-PKcs itself. Interestingly, these DNA-PKcs phosphorylation sites function in a distinct, and sometimes opposing manner, suggesting that they are differentially regulated via complex actions of both kinases and phosphatases. In this study we identified several phosphatase subunits as potential DSB-associated proteins. In particular, protein phosphatase 1 (PP1) is recruited to a DSB-mimicking substrate in Xenopusegg extracts and sites of laser microirradiation in human cells. Depletion of PP1 impairs NHEJ in both Xenopus egg extracts and human cells. PP1 binds multiple motifs of DNA-PKcs, regulates DNA-PKcs phosphorylation, and is required for DNA-PKcs activation after DNA damage. Interestingly, phosphatase 1 nuclear targeting subunit (PNUTS), an inhibitory regulator of PP1, is also recruited to DNA damage sites to promote NHEJ. PNUTS associates with the DNA-PK complex and is required for DNA-PKcs phosphorylation at Ser-2056 and Thr-2609. Thus, PNUTS and PP1 together finetune the dynamic phosphorylation of DNA-PKcs after DNA damage to mediate NHEJ.
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U2 - 10.1093/nar/gkx686
DO - 10.1093/nar/gkx686
M3 - Article
C2 - 28985363
AN - SCOPUS:85032948919
SN - 0305-1048
VL - 45
SP - 10583
EP - 10594
JO - Nucleic acids research
JF - Nucleic acids research
IS - 18
ER -