Human WRN Knockout Cell Line-HeLa
Cat.No. : CSC-RT1458
Host Cell: HeLa Target Gene: WRN
Size: 1x10^6 cells/vial, 1mL Validation: Sequencing
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Cell Line Information
Cell Culture Information
Safety and Packaging
| Cat. No. | CSC-RT1458 |
| Cell Line Information | A stable cell line with a homozygous knockout of human WRN using CRISPR/Cas9. |
| Target Gene | WRN |
| Host Cell | HeLa |
| Shipping | 10^6 cells/tube |
| Storage | Liquid nitrogen |
| Species | Human |
| Gene ID | 7486 |
| Revival | Rapidly thaw cells in a 37°C water bath. Transfer contents into a tube containing pre-warmed media. Centrifuge cells and seed into a 25 cm2 flask containing pre-warmed media. |
| Media Type | Cells were cultured in DMEM supplemented with 10% fetal bovine serum. |
| Growth Properties | Cells are cultured as a monolayer at 37°C in a humidified atmosphere with 5% CO2. Split at 80-90% confluence, approximately 1:4-1:6. |
| Freeze Medium | Complete medium supplemented with 10% (v/v) DMSO |
| Mycoplasma | Negative |
| Format | One frozen vial containing millions of cells |
| Storage | Liquid nitrogen |
| Safety Considerations |
The following safety precautions should be observed. 1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum. 2. No eating, drinking or smoking while handling the stable line. 3. Wash hands after handling the stable line and before leaving the lab. 4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells. 5. All waste should be considered hazardous. 6. Dispose of all liquid waste after each experiment and treat with bleach. |
| Ship | Dry ice |
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Background
Case Study
Applications
WRN, also known as the Werner syndrome gene, plays an important role in the fields of genetics and age-related diseases. The gene is located on chromosome 8 and encodes the Werner protein, which is involved in a variety of critical cellular processes, including DNA repair, replication, and maintaining telomere integrity. The WRN protein functions as a helicase and exonuclease, unwinding DNA and removing nucleotides from the ends of DNA strands, respectively. These dual enzymatic activities are essential for resolving complex DNA structures that arise during replication and repair. Through these mechanisms, WRN ensures that DNA is accurately replicated and errors or damage are corrected, thereby protecting genetic information across cell generations.
Mutations in the WRN gene are associated with Werner syndrome, an autosomal recessive disorder characterized by premature aging. People with Werner syndrome display common symptoms associated with aging, including cataracts, skin atrophy, osteoporosis, and an increased risk of cancer and cardiovascular disease, which typically appear in their third or fourth decade of life. Molecular studies have found that these mutations often result in truncated or nonfunctional WRN proteins, which impair DNA repair mechanisms and lead to genomic instability.
Mutations in the Werner (WRN) RECQL helicase are associated with premature aging syndrome (Werner syndrome, WS) and susceptibility to multiple cancers. In patients with solid cancers, WRN RECQL helicase deficiency is associated with improved overall survival after treatment with TOP1 inhibitors, which stabilize pathological TOP1-DNA-covalent-complexes (TOP1cc) on the genome. However, the potential mechanisms of WRN in conferring chemoresistance to TOP1 inhibitors remain unexplored. Here, genome-wide transcriptome analysis of ~25,000 genes revealed robust activation of NF-κB-dependent prosurvival genes in response to TOP1cc. CRISPR-Cas9 knockout, shRNA silencing, and underexpression of WRN render multiple cancers hypersensitive to TOP1 inhibitors. Researchers demonstrate that WRN orchestrates TOP1cc repair through both proteasome-dependent and proteasome-independent processes, unleashing robust ssDNA generation. This in turn provides signaling for CHK1-mediated NF-κB activation through IκBα-degradation and nuclear localization of p65 protein. Interestingly, site-directed mutagenesis and rescue experiments revealed that neither the RECQL helicase nor the DNA exonuclease enzymatic activities of WRN (WRNE84A, WRNK577M, and WRNE84A-K577M) are required for TOP1cc removal, ssDNA generation, and NF-κB activation signaling.
WRN knockout (WRN-KO) cells (with empty vector) were defective in removing TOP1cc compared to WRN-WT (Figure 1d, e). Ectopic expression of WRNWT in WRN-KO cells triggered complete removal of TOP1cc. Unexpectedly, ectopic expression of WRN single and double mutants for both exonuclease and helicase functions also almost completely rescued the phenotype of WRN-KO cells by enabling TOP1cc repair (Figure 1d, e). To further validate the redundant role of WRN exonuclease activity in TOP1cc removal, researchers generated the WRNΔExo deletion mutant (Figure 1b), which lacks the exonuclease domain (1-230 aa). Ectopic expression of WRNWT and WRNΔExo almost completely rescued the phenotype of WRN-KO cells, as TOP1cc was removed by CPT treatment. Together, these results suggest that non-enzymatic functions of WRN regulate TOP1cc removal by nanomolar CPT.
Figure 1. Non-enzymatic role of WRN in TOP1cc removal. (Gupta, Pooja, et al. 2022)
Cancer Research: WRN knockout HeLa cell lines are widely used in cancer research to understand the role of the WRN gene in tumorigenesis. Since HeLa cells are derived from cervical cancer, removing the WRN gene helps scientists study how this gene affects cancer cell proliferation, survival, and DNA repair mechanisms.
Genome Stability Studies: WRN is essential for maintaining genome stability. By creating a WRN knockout in HeLa cells, researchers can study how the loss of WRN affects chromosome integrity, potentially providing insights into diseases such as Werner syndrome.
Drug Screening: These cell lines serve as valuable models for high-throughput drug screening. Compounds that specifically target pathways or weaknesses in WRN-deficient cells can be identified, facilitating the development of treatments for cancer or diseases associated with WRN deficiency.
DNA Repair Mechanism Analysis: WRN knockout HeLa cells are essential for dissecting DNA repair pathways. Researchers use them to study how the loss of WRN affects various DNA repair processes, including homologous recombination and non-homologous end joining, providing insights into the molecular mechanisms of DNA repair.
Aging Research: Given that WRN has been implicated in premature aging diseases, WRN knockout HeLa cells could help researchers explore the cellular aging process.
For research use only. Not intended for any clinical use.
