|
|
GEO help: Mouse over screen elements for information. |
|
Status |
Public on May 08, 2021 |
Title |
Ku70 and Ligase IV deficiencies reveal distinct alternative end-joining outcomes in G1-arrested progenitor B cells |
Organism |
Mus musculus |
Experiment type |
Other
|
Summary |
Classical nonhomologous end-joining (C-NHEJ) repairs DNA double-stranded breaks (DSBs) throughout interphase but is thought to predominate in G1-phase when homologous recombination is unavailable. Complexes containing the Ku70/80 ("Ku") and XRCC4/Ligase IV (Lig4) core C-NHEJ factors are required, respectively, for sensing and joining DSBs. While such factors are exclusively required for joining RAG1/2-initiated DSBs during V(D)J recombination in G1-phase lymphocyte progenitors, cycling cells deficient for core C-NHEJ factors join chromosomal DSBs by alternative end-joining (A-EJ) pathways. Restriction of V(D)J recombination to C-NHEJ has been attributed to RAG-mediated exclusion of A-EJ; however, it remains unclear whether A-EJ is similarly excluded from more general DSBs in G1. Here, we report that Ku actively and robustly suppresses A-EJ of RAG1/2 and two classes of engineered endonuclease-mediated DSBs in G1-arrested progenitor B cell lines. Thus, while targeted DSBs remain as free broken ends in Lig4-deficient G1-arrested progenitor B cells, deletion of Ku70 in Lig4-deficient cells restores DSB rejoining and translocation to levels observed in Ku70-deficient counterparts. Correspondingly, while V(D)J recombination is abrogated in Ligase4-deficient lines, V(D)J-like joining occurs in Ku70-deficient and Ku70/Lig4 double-deficient lines through a translocation-based A-EJ mechanism. We conclude that in G1, Lig4-deficient progenitor B cells are functionally end-joining deficient due to a near complete Ku-dependent block in A-EJ. Thus, the differential impacts of Ku deficiency versus XRCC4/Ligase IV deficiency on V(D)J recombination, severity of neuronal apoptosis, and embryonic development, including Ku-deficiency rescuing Lig4-deficient embryonic lethality, may be explained by Ku-mediated inhibition of A-EJ in the G1 cell cycle phase.
|
|
|
Overall design |
We employed JoinT-seq, HTGTS-Rep-Rejoin and LAM-HTGTS to map genome-wide and rejoining repair outcome in G1-arrested murine pro-B cells and to elucidate the roles of Ku70 and Lig4 in repair pathway choice. We performed GRO-seq to study the gene transcription activity in cycling and G1-arrested v-Abl transformed pro-B cells.
|
|
|
Contributor(s) |
Liang Z, Kumar V, Le Bouteiller M, Zurita J, Kenrick J, Lin SG, Lou J, Hu J, Ye AY, Boboila C, Alt FW, Frock RL |
Citation(s) |
34006647 |
Submission date |
Dec 01, 2020 |
Last update date |
May 25, 2021 |
Contact name |
Marie Le Bouteiller |
E-mail(s) |
marie.lebouteiller@gmail.com
|
Organization name |
Stanford University
|
Department |
Radiation Oncology
|
Street address |
269 campus drive
|
City |
Stanford |
State/province |
CA |
ZIP/Postal code |
94305 |
Country |
USA |
|
|
Platforms (3) |
|
Samples (116)
|
|
Relations |
BioProject |
PRJNA681890 |
SRA |
SRP295405 |
Supplementary file |
Size |
Download |
File type/resource |
GSE162453_RAW.tar |
3.1 Gb |
(http)(custom) |
TAR (of BW, TXT) |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
|
|
|
|
|