Human TNFRSF1A Knockout Cell Line-A549
Cat.No. : CSC-RT2777
Host Cell: A549 Target Gene: TNFRSF1A
Size: 1x10^6 cells/vial, 1mL Validation: Sequencing
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Cell Line Information
Cell Culture Information
Safety and Packaging
| Cat. No. | CSC-RT2777 |
| Cell Line Information | This cell is a stable cell line with a homozygous knockout of human TNFRSF1A using CRISPR/Cas9. |
| Target Gene | TNFRSF1A |
| Host Cell | A549 |
| Size Form | 1 vial (>10^6 cell/vial) |
| Shipping | Dry ice package |
| Storage | Liquid Nitrogen |
| Species | Human |
| 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. |
| 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
The TNFRSF1A gene encodes the protein tumor necrosis factor receptor superfamily member 1A (TNFRSF1A), also known as tumor necrosis factor receptor 1 (TNFR1) or CD120a. The gene is located on human chromosome 12. As one of the main receptors for TNFα, it can activate multiple intracellular signaling pathways. One important pathway is the activation of the transcription factor NF-κB, which plays a key role in regulating immune responses and inflammation. In addition, TNFR1 is involved in mediating apoptosis (programmed cell death) and acts as a regulator of inflammation.
Mutations in the extracellular domain of the TNFRSF1A gene are associated with a genetic disorder called tumor necrosis factor receptor-associated periodic syndrome (TRAPS), also known as periodic fever syndrome. The disease is characterized by recurring fevers, abdominal pain, and muscle pain. The mechanism of the disease is thought to involve impaired clearance of the receptor, resulting in a prolonged and inappropriate inflammatory response. Studies have also shown that TNFRSF1A mutations can lead to an increased risk of developing multiple sclerosis (MS), an autoimmune disease that affects the central nervous system. Furthermore, higher TNFRSF1A serum levels have been observed in psychiatric disorders such as schizophrenia and bipolar disorder, which correlate with the severity of psychotic symptoms.
BRAF is a key mediator of RAS signaling, with activating mutations present in approximately 50% of melanoma patients. Pharmacological inhibition of BRAF or the downstream MAP kinase MEK is highly effective in treating BRAF mutant melanoma. Studies here demonstrate that treatment of melanoma patients with BRAF and MEK inhibitors (MEKi) activates tumor NF-κB activity. In melanoma and lung cancer cells, MEKi increases cell surface expression of TNFα receptor 1 (TNFR1, also known as tumor necrosis factor receptor superfamily member 1A (TNFRSF1A)), thereby enhancing NF-κB activation and increasing expression of genes regulated by TNFα and IFNγ. Screening of 289 targeted agents for their ability to increase expression of TNFα and IFNγ target genes suggests that this is a general activity of MEK and ERK kinase inhibitors. Treatment with MEKi resulted in a new susceptibility of lung cancer cells to TNFα- and IFNγ-induced apoptosis, whereas these cells were resistant to MEKi killing and had increased cell cycle arrest. Ablation of TNFR1 expression on lung cancer cells impaired the antitumor efficacy of MEKi, whereas administration of TNFα and IFNγ to MEKi-treated mice enhanced the antitumor response. These findings define a novel cytokine response-regulating function of MEKi that could be exploited therapeutically.
Here, the researchers observed that the growth inhibitory effect of MEKi was partially attenuated in TNFR1 knockout (TNFR1KO) A549 cells, but re-expression of TNFR1 in these cells resensitized them to MEKi (Figure 1A). These results suggest that activation of autocrine TNFR1/TNFα signaling by MEKi can enhance growth inhibition. Next, they tested the effects of exogenous addition of TNFα and IFNγ. TNFα alone and TNFα + IFNγ were found to reduce the number of viable cells modestly. MEKi reduced the number of cells. However, the combination of MEKi with TNFα + IFNγ resulted in the greatest reduction in the number of viable cells (Figure 1B). Treatment with MEKi in the presence of both cytokines resulted in the greatest percentage of cells in G1 phase, the lowest percentage of cells in S phase (Figure 1C), and the highest activation of the apoptotic marker cleaved caspase-3 (Figure 1D).
Figure 1. TNFα and IFNγ enhance MEKi-induced growth suppression and cell death. (Xie M, et al., 2019)
Utilizing TNFR1 knockout A549 cells, researchers next tested the in vivo role of TNFR1 signaling in MEKi antitumor response in an orthotopic lung tumor model using immunodeficient SCID (Prkdcscid) and beige (Lystbg) mice. Unlike mice treated with vehicle, administration of trametinib to mice was associated with minimal tumor burden (Figure 1E and F). In contrast, no reduction in tumor burden was observed after MEKi treatment in mice bearing TNFR1 KO tumors (Figure 1E and F). Importantly, pERK was strongly inhibited by MEKi in KO tumors, indicating that trametinib retains MEK targeting ability in these tumors (Figure 1G). Combined with the in vitro findings, these results suggest that TNFR1 signaling plays a key role in the MEKi antitumor response.
Cancer Research: TNFRSF1A knockout in the A549 cell line can be used to study the role of TNF receptors in non-small cell lung cancer (NSCLC). This model helps understand how loss of TNFRSF1A affects tumor growth, metastasis, and response to therapy, paving the way for new targeted therapies.
Inflammatory Response Research: TNFRSF1A is essential in mediating inflammatory responses. Using this knockout cell line, researchers can study the molecular pathways of inflammation and the specific role of TNFRSF1A in inflammatory diseases such as rheumatoid arthritis and Crohn's disease.
Drug Discovery and Testing: This cell line can serve as a model system for screening potential therapeutic agents targeting the TNF pathway. By using the TNFRSF1A knockout model, researchers can identify drugs that effectively modulate the TNF signaling cascade, thereby developing new anti-inflammatory and anti-cancer drugs.
Signal Transduction Research: Researchers can explore how loss of TNFRSF1A affects downstream signaling events such as apoptosis, cell survival, and cytokine production, gaining insight into complex cell signaling networks.
Immunology Research: Given the role of TNFRSF1A in immune regulation, this knockout cell line is a valuable tool for immunologists. It helps study how TNFRSF1A affects immune cell responses and aids in the development of immunotherapies and vaccines for a variety of diseases, including autoimmune diseases and cancer.
For research use only. Not intended for any clinical use.
