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| Status |
Public on Sep 18, 2023 |
| Title |
Genome-wide analysis of DNA-PK-bound MRN cleavage products supports a sequential model of DSB repair pathway choice |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
The Mre11-Rad50-Nbs1 (MRN) complex recognizes and processes DNA double-strand breaks for homologous recombination by performing short-range removal of 5ʹ strands. Endonucleolytic processing by MRN requires a stably bound protein at the break site—a role we postulate is played by DNA-dependent protein kinase (DNA-PK) in mammals. Here we interrogate the sites of MRN-dependent processing by isolating and sequencing DNA-PK-bound DNA fragments that are products of MRN cleavage. These intermediates are generated with highest efficiency when DNA-PK is catalytically blocked, yielding products within 200 bp of the break site, whereas DNA-PK products in the absence of kinase inhibition show much greater dispersal. Use of light-activated Cas9 to induce breaks facilitates temporal resolution of DNA-PK and Mre11 binding, showing that Mre11 and DNA-PK both bind to DNA ends before release of DNA-PK-bound products. These results support a sequential model of double-strand break repair involving collaborative interactions between homologous and non-homologous repair complexes.
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| Overall design |
Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for phospho-DNA-PKcs(S2056), GLASS-ChIP DNA sequencing for phospho-DNA-PKcs(S2056), and Mre11 ChIPseq
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| Contributor(s) |
Deshpande R, Marin-Gonzalez A, Ha T, Paull T, Barnes H, Woolley P |
| Citation(s) |
37717054 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R01 GM138548 |
DNA end processing by the Mre11/Rad50/Nbs1 complex in human cells |
UNIVERSITY OF TEXAS AUSTIN |
TANYA T PAULL |
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| Submission date |
Nov 22, 2022 |
| Last update date |
Sep 27, 2023 |
| Contact name |
Tanya T. Paull |
| E-mail(s) |
tpaull@utexas.edu
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| Phone |
5122327803
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| Organization name |
Univ. of Texas at Austin
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| Department |
Molecular Biosciences
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| Lab |
Paull
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| Street address |
2500 Speedway MBB 2.448
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| City |
Austin |
| State/province |
TX |
| ZIP/Postal code |
78712 |
| Country |
USA |
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| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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| Samples (58)
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| GSM6752995 |
GC, no 4-OHT, +NU-7441, +antibody, rep1 |
| GSM6752996 |
GC, no 4-OHT, +NU-7441, -antibody, rep1 |
| GSM6752997 |
GC, no 4-OHT, +NU-7441, +antibody, rep2 |
| GSM6752998 |
GC, plus 4-OHT, +NU-7441, +antibody, rep1 |
| GSM6752999 |
GC, plus 4-OHT, +NU-7441, -antibody, rep1 |
| GSM6753000 |
GC, plus 4-OHT, +NU-7441, +antibody, rep2 |
| GSM6753001 |
GC, no 4-OHT, +antibody, rep1 |
| GSM6753002 |
GC, no 4-OHT, +antibody, rep2 |
| GSM6753003 |
GC, no 4-OHT, -antibody, rep1 |
| GSM6753004 |
GC, no 4-OHT, -antibody, rep2 |
| GSM6753005 |
GC, plus 4-OHT, +antibody, rep1 |
| GSM6753006 |
GC, plus 4-OHT, +antibody, rep2 |
| GSM6753007 |
GC, plus 4-OHT, -antibody, rep1 |
| GSM6753008 |
GC, plus 4-OHT, -antibody, rep2 |
| GSM6753009 |
PC, no 4-OHT, +NU-7441, +antibody |
| GSM6753010 |
PC, no 4-OHT, +NU-7441, input |
| GSM6753011 |
PC, plus 4-OHT, +NU-7441, +antibody |
| GSM6753012 |
PC, plus 4-OHT, +NU-7441, input |
| GSM6753013 |
PC, no 4-OHT, +antibody |
| GSM6753014 |
PC, no 4-OHT, input |
| GSM6753015 |
PC, plus 4-OHT, +antibody |
| GSM6753016 |
PC, plus 4-OHT, input |
| GSM6753017 |
GC, no light, +NU-7441, +antibody, rep1 |
| GSM6753018 |
GC, no light, +NU-7441, +antibody, rep2 |
| GSM6753019 |
GC, 15m post-light, +NU-7441, +antibody, rep1 |
| GSM6753020 |
GC, 15m post-light, +NU-7441, +antibody, rep2 |
| GSM6753021 |
GC, 60m post-light, +NU-7441, +antibody, rep1 |
| GSM6753022 |
GC, 60m post-light, +NU-7441, +antibody, rep2 |
| GSM6753023 |
PC, no light, +NU-7441, +antibody, rep1 |
| GSM6753024 |
PC, no light, +NU-7441, +antibody, rep2 |
| GSM6753025 |
PC, 15m post-light, +NU-7441, +antibody, rep1 |
| GSM6753026 |
PC, 15m post-light, +NU-7441, +antibody, rep2 |
| GSM6753027 |
PC, 60m post-light, +NU-7441, +antibody, rep1 |
| GSM6753028 |
PC, 60m post-light, +NU-7441, +antibody, rep2 |
| GSM6753029 |
Mre11, no light, +NU-7441, +antibody, rep1 |
| GSM6753030 |
Mre11, no light, +NU-7441, +antibody, rep2 |
| GSM6753031 |
Mre11, 15m post-light, +NU-7441, +antibody, rep1 |
| GSM6753032 |
Mre11, 15m post-light, +NU-7441, +antibody, rep2 |
| GSM6753033 |
Mre11, 60m post-light, +NU-7441, +antibody, rep1 |
| GSM6753034 |
Mre11, 60m post-light, +NU-7441, +antibody, rep2 |
| GSM6756896 |
GC, plus 4-OHT, +NU-7441, 1 hr only, +antibody |
| GSM6756897 |
GC, plus 4-OHT, +NU-7441, 1 hr only, -antibody |
| GSM6756898 |
GC, plus 4-OHT, +NU-7441, + PFM01, 1 hr only, +antibody |
| GSM6756899 |
GC, plus 4-OHT, +NU-7441, + PFM01, 1 hr only, -antibody |
| GSM6756900 |
GC, plus 4-OHT, +NU-7441, + PFM39, 1 hr only, +antibody |
| GSM6756901 |
GC, plus 4-OHT, +NU-7441, + PFM39, 1 hr only, -antibody |
| GSM7498202 |
CtIP GC, no 4-OHT, +antibody, rep1 |
| GSM7498203 |
CtIP GC, no 4-OHT, +antibody, rep2 |
| GSM7498204 |
CtIP GC, no 4-OHT, -antibody |
| GSM7498205 |
CtIP GC, plus 4-OHT, +antibody, rep1 |
| GSM7498206 |
CtIP GC, plus 4-OHT, +antibody, rep2 |
| GSM7498207 |
CtIP GC, plus 4-OHT, -antibody |
| GSM7498208 |
CtIP PC, no 4-OHT, +antibody, rep1 |
| GSM7498209 |
CtIP PC, no 4-OHT, +antibody, rep2 |
| GSM7498210 |
CtIP PC, no 4-OHT, input |
| GSM7498211 |
CtIP PC, plus 4-OHT, +antibody, rep1 |
| GSM7498212 |
CtIP PC, plus 4-OHT, +antibody, rep2 |
| GSM7498213 |
CtIP PC, plus 4-OHT, input |
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| Relations |
| BioProject |
PRJNA904374 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE218590_CtIP_GC_no_4-OHT_+antibody.bigwig |
45.8 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_CtIP_GC_plus_4-OHT_+antibody.bigwig |
40.1 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_CtIP_PC_no_4-OHT_+antibody.bigwig |
656.5 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_CtIP_PC_plus_4-OHT_+antibody.bigwig |
718.3 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC+DNAPKiplus4H1hronly+PFM01+antibody.bigwig |
70.0 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC+DNAPKiplus4H1hronly+PFM39+antibody.bigwig |
133.9 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC+DNAPKiplus4H1hronly+antibody.bigwig |
124.3 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_+DNA-PKi_15m_post-light_+antibody_R1_and_R2.bigwig |
67.2 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_+DNA-PKi_60m_post-light_+antibody_R1_and_R2.bigwig |
74.0 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_+DNA-PKi_no_light_+antibody_R1_and_R2.bigwig |
59.0 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_+DNAPKi_no_4H_+antibody__R1_and_R2.bigwig |
417.0 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_+DNAPKi_plus_4H_+antibody__R1_and_R2.bigwig |
335.2 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_no_4H_+antibody_R1_and_R2.bigwig |
301.8 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_GC_plus_4H_+antibody_R1_and_R2.bigwig |
344.7 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_Mre11_+DNA-PKi_15m_post-light_+antibody_R1_and_R2.bigwig |
176.1 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_Mre11_+DNA-PKi_60m_post-light_+antibody_R1_and_R2.bigwig |
155.1 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_Mre11_+DNA-PKi_no_light_+antibody_R1_and_R2.bigwig |
176.5 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_+DNA-PKi_15m_post-light_+antibody_R1_and_R2.bigwig |
353.9 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_+DNA-PKi_60m_post-light_+antibody_R1_and_R2.bigwig |
343.4 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_+DNA-PKi_no_light_+antibody_R1_and_R2.bigwig |
318.6 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_+DNAPKi_no_4H_+antibody.bigwig |
319.0 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_+DNAPKi_plus_4H_+antibody.bigwig |
264.2 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_no_4H_+antibody.bigwig |
363.0 Mb |
(ftp)(http) |
BIGWIG |
| GSE218590_PC_plus_4H_+antibody.bigwig |
383.7 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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