Stable CDK12 knock-out ovarian cancer cells do not show an increased sensitivity to cisplatin and PARP inhibitor treatment
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA758916 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR15669459.fastq.gz | Fastqsanger.gz | |||
| SRR15669460.fastq.gz | Fastqsanger.gz | |||
| SRR15669461.fastq.gz | Fastqsanger.gz | |||
| SRR15669462.fastq.gz | Fastqsanger.gz | |||
| SRR15669463.fastq.gz | Fastqsanger.gz |
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Project description:We stabily knock-out CDK12 in A2780 cell line and performed RNA-Seq profiling comparing WT and KO conditions.
2022-07-11 | GSE183041 | GEO
Project description:Cyclin-dependent kinase 12 (CDK12) is a serine/threonine kinase involved in the regulation of RNA polymerase II and in the transcription of a subset of genes involved in the DNA damage response. CDK12 is one of the most mutated genes in ovarian carcinoma. These mutations result in loss-of-function and can predict the responses to PARP1/2 inhibitor and platinum. To investigate the role of CDK12 in ovarian cancer, CRISPR/Cas9 technology was used to generate a stable CDK12 knockout (KO) clone in A2780 ovarian carcinoma cells. This is the first report on a CDK12 null cell line. The clone had slower cell growth and was less clonogenic than parental cells. These data were confirmed in vivo, where CDK12 KO transplanted cells had a much longer time lag and slightly slower growth rate than CDK12-expressing cells. The slower growth was associated with a higher basal level of apoptosis, but there were no differences in the basal level of autophagy and senescence. While cell cycle distribution was similar in parental and knockout cells, there was a doubling in DNA content, with an almost double modal number of chromosomes in the CDK12 KO clone which, however did not display any increase in γH2AX, a marker of DNA damage. We found partial down-regulation of the expression of DNA repair genes at the mRNA level and, among the down-regulated genes, an enrichment in the G2/M checkpoint genes. Although the biological features of CDK12 KO cells are compatible with the function of CDK12, contrary to some reports, we could not find any difference in the sensitivity to cisplatin and olaparib between wild-type and CDK12 KO cells.
| S-EPMC9328802 | biostudies-literature
Project description:We describe the epigenetic profiling of the H3K9me2 and HP1a in Drosophila third instar larvae before and after CDK12 depletion by RNA interference (RNAi). Here we show that CDK12 regulates heterochromatin dynamics in Drosophila chromosomes. Depletion of CDK12 induces the increased HP1a and H3K9me2 binding profile on the coding region of euchromatic genes, with the X chromosome being the most affected. These results are consistent with the polytene chromosome immunostaining pattern of HP1a and H3K9me2 after CDK12 knockdown in our initial cytological observations, which show that CDK12 depletion induce heterochromatin spreading on euchromatic arms, especially on the X chromosome. This study describes a novel role of the CDK12 complex in controlling the epigenetic transition between euchromatin and heterochromatin. Examination of the genome-wide H3K9me2 and HP1a binding profile in wildtype larvae (WT) and CDK12-depleted larvae (CDK12-KD). Examination of the genome-wide CDK12 binding profile in wildtype larvae (WT). Twelve independent immunoprecipitations were conducted for each antibody. Two biological replicates were performed.
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Project description:In a forward genetic screen, we have previously identified a null mutant of Cdk12 that results in alterations in actin dynamics, the axon initial segment and electrophysiology in Drosophila melanogaster. To decipher how Cdk12 may be having these effects, we extracted RNA from pooled Drosophila heads and compared Cdk12-null mutants to controls at the transcriptome level.
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Project description:Current treatment strategies for advanced prostate cancer (PCa) primarily rely on androgen receptor (AR)-directed therapies. However, the emergence of castration-resistant prostate cancer (CRPC) and resistance to AR signaling inhibitors (ARSI) pose substantial challenges. This study introduces a triple degrader, BSJ-5-63, targeting CDK12, CDK7, and CDK9, which has the potential to transform CRPC therapy. BSJ-5-63 uniquely downregulates homologous recombination repair (HRR) genes, including BRCA1 and BRCA2, by degrading CDK12, and it also attenuates AR signaling by degrading CDK7 and CDK9, further enhancing BSJ-5-63's therapeutic impact. Importantly, BSJ-5-63 induces a 'BRCAness' state that persists for a significant duration, enabling rational combination therapy with PARP inhibitors (PARPis) while minimizing drug-related toxicity and resistance. In both in vitro and in vivo studies, potent anti-proliferative effects were observed in both AR-positive and -negative CRPC cells. This research presents a promising multi-pronged approach for CRPC treatment, addressing both DNA repair mechanisms and AR signaling, with the potential to benefit a wide range of patients, regardless of their BRCA1/2 mutational status.
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Project description:Mutations that attenuate DNA repair by homologous recombination (HR) promote tumorigenesis and sensitize cells to chemotherapeutic agents that cause replication fork collapse, a phenotype known as “BRCAness.” BRCAness tumors arise from loss-of-function mutations in 22 genes. Of these genes, all but one (Cdk12) directly function in the HR repair pathway. Cdk12 phosphorylates Serine 2 of the RNA Polymerase II (RNAPII) C-terminal domain (CTD) heptapeptide repeat, a modification that regulates transcription elongation, splicing, and cleavage/polyadenylation. Genome-wide expression studies suggest that Cdk12 depletion abrogates the expression of several HR genes relatively specifically, blunting HR repair. This observation suggests that Cdk12 mutational status may predict tumor sensitivity to targeted treatments against BRCAness, such as Parp 1 inhibitors, and that small-molecule inhibitors of Cdk12 may induce sensitization of otherwise HR-competent tumors to these treatments. Despite this growing clinical interest, the mechanism behind the apparent specificity of Cdk12 in regulating HR genes remains unknown. Here we find that Cdk12 globally suppresses intronic polyadenylation events, enabling the production of full-length gene products. Many HR genes harbor significantly more intronic polyadenylation sites compared to all expressed genes, and the cumulative effect of these sites accounts for the increased sensitivity of HR gene expression to Cdk12 loss. Finally, we find evidence that Cdk12 loss-of-function mutations cause increased intronic polyadenylation within HR genes in human tumors, suggesting that this mechanism is conserved. This work clarifies the biological function of CDK12 and underscores its potential both as a chemotherapeutic target and as a tumor biomarker.
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Project description:PARP inhibitor and platinum based drugs such as cisplatin are promising therapies for triple negative breast cancer and exploit the deficiencies in BRCA1 or BRCA2, or homologous recombination repair defects. However, PARP inhibitor resistance is proven to be a major clinical problem. Acquired PARP inhibitor resistance has been linked with co-resistance to platinum-based drugs. To determine how acquired olaparib resistance affects cisplatin response and whether this is influenced by their BRCA1 status, we performed RNAseq transcriptome analysis of isogenic triple negative breast cancer models of olaparib resistance with normal and mutant BRCA1.
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Project description:We describe the epigenetic profiling of the H3K9me2 and HP1a in Drosophila third instar larvae before and after CDK12 depletion by RNA interference (RNAi). Here we show that CDK12 regulates heterochromatin dynamics in Drosophila chromosomes. Depletion of CDK12 induces the increased HP1a and H3K9me2 binding profile on the coding region of euchromatic genes, with the X chromosome being the most affected. These results are consistent with the polytene chromosome immunostaining pattern of HP1a and H3K9me2 after CDK12 knockdown in our initial cytological observations, which show that CDK12 depletion induce heterochromatin spreading on euchromatic arms, especially on the X chromosome. This study describes a novel role of the CDK12 complex in controlling the epigenetic transition between euchromatin and heterochromatin.
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Project description:PARP-6, a member of a family of enzymes (17 in humans) known as poly-ADP- ribose polymerases (PARPs), is a neuronally enriched PARP. While previous studies from our group show that PARP-6 is a regulator of dendrite morphogenesis in hippocampal neurons, its function in the nervous system in vivo is poorly understood. Here, we describe the generation of a PARP-6 loss-of-function mouse model for examining the function of PARP-6 during neurodevelopment in vivo. Using CRISPR-Cas9 mutagenesis, we generated a mouse line that expresses a PARP-6 truncated variant (PARP-6TR) in place of PARP-6WT. Unlike PARP-6WT, PARP-6TR is devoid of catalytic activity. Homozygous PARP-6TR do not exhibit obvious neuromorphological defects during development, but nevertheless die perinatally. This suggests that PARP-6 catalytic activity is important for postnatal survival. We also report PARP-6 mutations in six patients with several neurodevelopmental disorders, including microencephaly, intellectual disabilities, and epilepsy. The most severe mutation in PARP-6 (C563R) results in the loss of catalytic activity. Expression of the PARP-6C563R mutant in hippocampal neurons decreases dendrite morphogeneis. Taken together, these results suggest that PARP-6 is an essential gene in mice, and the loss of PARP-6 catalytic activity has detrimental effects on neuronal function in humans.
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Project description:Determining interaction partners with CDK12 using IP-MS. Capturing global proteome expression changes as a result of CDK12 knockdown.
2017-03-21 | PXD004184 | Pride