|
| Status |
Public on Jun 19, 2024 |
| Title |
CDK12 prevents MYC-induced transcription-replication conflicts [EdU-seq] |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
|
| Summary |
In order to identify genes and pathways necessary to preserve genome integrity upon Myc over-expression, we conducted a large siRNA-based screen in isogenic lines. We discovered several genes that suppressed the Myc-induced DNA-damage response and that were essential for the cell survival upon Myc activation. We idntified CDK12, a cyclin dependent kinase involved in transcriptional control and genome stability. We uncovered a novel and unexpected role of CDK12 in controlling transcription at loci proximal to DNA damaged sites and dissected its upstream regulatory pathways and downstream effectors. Mechanistic studies and genome-wide mapping of replication dynamics and DSBs revealed how CDK12 is essential to suppress intrinsic transcription-replication conflicts, thus avoiding cytotoxic DNA damage in cancer cells. Overall, this study uncovers a novel role for CDK12 and a new liability of Myc-driven cancers, which could be exploited for therapeutic purposes.
|
| |
|
| Overall design |
U2OS MycER cells (U20S cells expressing the MycER chimera) were infected with virus expressing non-targeting shRNAs (shNT) or shRNA targeting CDK12 (shCDK12). ShRNA expression was activated with 1 µg/mL doxycycline, while MycER was activated with 300 nM OHT for 40h. Cells were then treated with 100 ng/ml nocodazole (Sigma, Cat. No. SML-1665) for 8h to induce mitotic arrest. G2/M arrested cells were isolated by shake-off, washed with PBS and released in warm medium containing 25 μM EdU (Invitrogen, Cat. No. A10044) and 2 mM hydroxyurea (Sigma, Cat. No. H8627-5G). After 24 h, cells were collected and fixed in 90% methanol.
|
| |
|
| Contributor(s) |
Campaner S, Curti L, Croci O, Bianchi N |
| Citation(s) |
39155303 |
| |
| Submission date |
Jul 05, 2023 |
| Last update date |
Sep 23, 2024 |
| Contact name |
Stefano Campaner |
| E-mail(s) |
stefano.campaner@iit.it
|
| Organization name |
fondazione Istituto italiano di tecnologia
|
| Department |
Center for Genomic Science
|
| Lab |
Cancer Biology
|
| Street address |
Via adamello 16
|
| City |
Milano |
| ZIP/Postal code |
20139 |
| Country |
Italy |
| |
|
| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
|
| Samples (12)
|
|
| This SubSeries is part of SuperSeries: |
| GSE236553 |
CDK12 prevents MYC-induced transcription-replication conflicts |
|
| Relations |
| BioProject |
PRJNA991567 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE236550_EduSeq_shCDK12_HU_merge.bed.gz |
97.6 Kb |
(ftp)(http) |
BED |
| GSE236550_EduSeq_shCDK12_HU_merge.bw |
126.1 Mb |
(ftp)(http) |
BW |
| GSE236550_EduSeq_shCDK12_OHT_HU_merge.bed.gz |
96.5 Kb |
(ftp)(http) |
BED |
| GSE236550_EduSeq_shCDK12_OHT_HU_merge.bw |
182.8 Mb |
(ftp)(http) |
BW |
| GSE236550_EduSeq_shNT_HU_merge.bed.gz |
78.4 Kb |
(ftp)(http) |
BED |
| GSE236550_EduSeq_shNT_HU_merge.bw |
121.8 Mb |
(ftp)(http) |
BW |
| GSE236550_EduSeq_shNT_OHT_HU_merge.bed.gz |
94.4 Kb |
(ftp)(http) |
BED |
| GSE236550_EduSeq_shNT_OHT_HU_merge.bw |
181.4 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
Less...
More...