scholarly journals Phosphorylation in the serine/threonine 2609–2647 cluster promotes but is not essential for DNA-dependent protein kinase-mediated nonhomologous end joining in human whole-cell extracts

2007 ◽  
Vol 35 (12) ◽  
pp. 3869-3878 ◽  
Author(s):  
Lawrence F. Povirk ◽  
Rui-Zhe Zhou ◽  
Dale A. Ramsden ◽  
Susan P. Lees-Miller ◽  
Kristoffer Valerie
Keyword(s):  
Protein Kinase ◽  
Nonhomologous End Joining ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Whole Cell ◽  
Cell Extracts ◽  
Dependent Protein

DNA end sequestration by DNA-dependent protein kinase and end joining of sterically constrained substrates in whole-cell extracts

2003 ◽  
Vol 42 (4) ◽  
pp. 279-287 ◽  
Author(s):  
Jae Wan Lee ◽  
Kedar V. Inamdar ◽  
Michele F. Hannah ◽  
Susan P. Lees-Miller ◽  
Lawrence F. Povirk
Keyword(s):  
Protein Kinase ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Whole Cell ◽  
Cell Extracts ◽  
Dependent Protein

scholarly journals DNA-dependent protein kinase in nonhomologous end joining: a lock with multiple keys?

2007 ◽  
Vol 179 (2) ◽  
pp. 183-186 ◽  
Author(s):  
Eric Weterings ◽  
David J. Chen
Keyword(s):  
Protein Kinase ◽  
Strand Break ◽  
Nonhomologous End Joining ◽  
End Joining ◽  
Dsb Repair ◽  
Dependent Protein Kinase ◽  
Dna Molecule ◽  
Dna Double Strand Break ◽  
Dependent Protein ◽  
Multiple Keys

The DNA-dependent protein kinase (DNA-PK) is one of the central enzymes involved in DNA double-strand break (DSB) repair. It facilitates proper alignment of the two ends of the broken DNA molecule and coordinates access of other factors to the repair complex. We discuss the latest findings on DNA-PK phosphorylation and offer a working model for the regulation of DNA-PK during DSB repair.

scholarly journals DNA-dependent protein kinase promotes DNA end processing by MRN and CtIP

2020 ◽  
Vol 6 (2) ◽  
pp. eaay0922 ◽  
Author(s):  
Rajashree A. Deshpande ◽  
Logan R. Myler ◽  
Michael M. Soniat ◽  
Nodar Makharashvili ◽  
Linda Lee ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Homologous Recombination ◽  
Single Molecule ◽  
Strand Break ◽  
Nonhomologous End Joining ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Mrn Complex ◽  
Dependent Protein ◽  
Dna Ends

The repair of DNA double-strand breaks occurs through nonhomologous end joining or homologous recombination in vertebrate cells—a choice that is thought to be decided by a competition between DNA-dependent protein kinase (DNA-PK) and the Mre11/Rad50/Nbs1 (MRN) complex but is not well understood. Using ensemble biochemistry and single-molecule approaches, here, we show that the MRN complex is dependent on DNA-PK and phosphorylated CtIP to perform efficient processing and resection of DNA ends in physiological conditions, thus eliminating the competition model. Endonucleolytic removal of DNA-PK–bound DNA ends is also observed at double-strand break sites in human cells. The involvement of DNA-PK in MRN-mediated end processing promotes an efficient and sequential transition from nonhomologous end joining to homologous recombination by facilitating DNA-PK removal.

scholarly journals DNA-dependent Protein Kinase Activity Is Not Required for Immunoglobulin Class Switching

2002 ◽  
Vol 196 (11) ◽  
pp. 1483-1495 ◽  
Author(s):  
Gayle C. Bosma ◽  
Jiyoon Kim ◽  
Teresa Urich ◽  
Donna M. Fath ◽  
Maria G. Cotticelli ◽  
...  
Keyword(s):  
B Cells ◽  
Protein Kinase ◽  
Scid Mice ◽  
Nonhomologous End Joining ◽  
Protein Kinase Activity ◽  
Dna Double Strand Breaks ◽  
Dna Breaks ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Dependent Protein

Class switch recombination (CSR), similar to V(D)J recombination, is thought to involve DNA double strand breaks and repair by the nonhomologous end–joining pathway. A key component of this pathway is DNA-dependent protein kinase (DNA-PK), consisting of a catalytic subunit (DNA-PKcs) and a DNA-binding heterodimer (Ku70/80). To test whether DNA-PKcs activity is essential for CSR, we examined whether IgM+ B cells from scid mice with site-directed H and L chain transgenes were able to undergo CSR. Although B cells from these mice were shown to lack DNA-PKcs activity, they were able to switch from IgM to IgG or IgA with close to the same efficiency as B cells from control transgenic and nontransgenic scid/+ mice, heterozygous for the scid mutation. We conclude that CSR, unlike V(D)J recombination, can readily occur in the absence of DNA-PKcs activity. We suggest nonhomologous end joining may not be the (primary or only) mechanism used to repair DNA breaks during CSR.

scholarly journals Nonhomologous end-joining deficiency of L5178Y-S cells is not associated with mutation in the ABCDE autophosphorylation cluster.

2006 ◽  
Vol 53 (1) ◽  
pp. 233-236 ◽  
Author(s):  
Kamil Brzóska ◽  
Marcin Kruszewski ◽  
Irena Szumiel
Keyword(s):  
Protein Kinase ◽  
Cell Line ◽  
In Vitro Test ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Cell Extracts ◽  
Dependent Protein ◽  
Phenotypic Features ◽  
Repair Defect

Cells with mutated autophosphorylation sites in the ABCDE cluster of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) are defective in the repair of ionising radiation-induced DSB, but show in an in vitro test the same DNA-PK activity as the cells possessing wild type enzyme. Nevertheless, the mutated DNA-PK is able to undergo ATP-dependent autophosphorylation and inactivation. This characteristics correspond well with the phenotypic features of the L5178Y-S (LY-S) cell line that is defective in DSB repair, shows a pronounced G1 phase radiosensitivity, but in which the level of DNA-PK activity present in total cell extracts is similar to that of its radioresistant counterpart L5178Y-R (LY-R) cell line. The purpose of this work was to examine the possible alterations in the sequence encoding the cluster of autophosphorylation sites in the DNA-dependent protein kinase in LY-S cells. Despite the presence of phenotypic features indicating the possibility of such alterations, no differences were found between the sequences coding for the autophosphorylation sites in L5178Y-R and L5178Y-S cells. In conclusion, the repair defect in LY-S cells is not related to the structure of the DNA-PK autophosphorylation sites (ABCDE casette).

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