Human TLR3 NF-kB/SEAP - (SEAPorter™) Stable Reporter Cell Line

Product Discontinued

Human TLR3 NF-kB/SEAP - (SEAPorter™) Stable Reporter Cell Line Summary

• Description: The TLR3 reporter line can be used for TLR3-dependent functional assays as well as screening of TLR3 agonists or antagonists. The TLR3 reporter cell line has been validated by flow cytometry (Fig. 1) and ligand dose response assay (Fig. 2).
  
  Contents: 3~4 x 10^6 cells
  Biosafety Level: 1
• Immunogen: The TLR3 reporter cell line is a stably co-transfected cell line which expresses full-length human Toll-like receptor 3 (TLR3) and the secreted alkaline phosphatase (SEAP) reporter gene under the transcriptional control of an NF-kB response element.
• Target Species: Human
• Specificity: TLR3/NF-kB/SEAP
• Selection Agent: Blasticidin and G418.
• Growth Properties: Adherent Morphology: Epithelial
• RCL Type: NF-kB/SEAP - (SEAPorter™)
• Host: HEK293
• Gene: TLR3

Applications/Dilutions

• Dilutions:
  • Flow Cytometry
  • Ligand Activation
• Agonist:
  • Polyinosinic-polycytidylic acid HMW NBP2-25288
• Readout System:
  • Secreted Alkaline Phosphatase Reporter Assay Kit (Colorimetric) NBP2-25285

Packaging, Storage & Formulations

• Storage: Store in gas phase of liquid nitrogen.
• Reconstitution Instructions: Growth Medium: DMEM with 4.5 g/L glucose + 10% FBS + 4 mM L-glutamine + 1 mM sodium pyruvate + 100 units/ml penicillin + 0.1 mg/ml streptomycin + 10 ug/ml blasticidin + 0.5 mg/ml G418 (Geneticin).

Details for Array

• Type: Cell

Notes

Assume all cultures are hazardous since they may harbor latent viruses or other organisms that are uncharacterized. The following safety precautions should be observed.
- Use pipette aids to prevent ingestion and keep aerosols down to a minimum.
- No eating, drinking or smoking while handling the TLR3 reporter cell line.
- Wash hands after handling the reporter cell line and before leaving the lab.
- Decontaminate work surface with disinfectant or 70% ethanol before and after working with cells.
- All waste should be considered hazardous.
- Dispose of all liquid waste after each experiment and treat with bleach.

Alternate Names for Human TLR3 NF-kB/SEAP - (SEAPorter™) Stable Reporter Cell Line

• CD283 antigen
• CD283
• TLR3
• toll-like receptor 3

Background

Toll-like receptor 3 (TLR3) is a type I transmembrane glycoprotein that contributes to the innate immune response, recognizing distinct pathogen-associate molecular patterns (PAMPs) and damage-associate molecular patterns (DAMPs) (1,2). The TLR family member TLR3 specifically recognizes and binds double-stranded RNA (dsRNA) from viruses and the synthetic analog polyriboinosinic:polyribocytidylic acid (poly(I:C)) (1-5). TLR3 is typically expressed in the endosomes of innate immune cells including macrophages, natural killer (NK) cells, and dendritic cells (DCs) (1-5). TLR3 is also localized on the cell surface of fibroblasts, epithelial cells, and vascular endothelial cells (1-5). The human TLR3 protein is 904 amino acids (aa) in length with a theoretical molecular weight (MW) of 104 kDa (6). It consists of a 23 aa signal sequence, a horseshoe-shaped 681 aa extracellular domain (ECD) containing 23 leucine-rich repeats (LRRs), a 21 aa helical transmembrane domain, and a 179 aa cytoplasmic region containing a Toll/IL-1 receptor (TIR) domain (1,6). Upon ligand binding, TLR3-ECD dimerizes and the adapter protein TIR-domain-containing adapter inducing interferon-beta (TRIF/TICAM1) is recruited (1-5). TRIF interacts with tumor necrosis factor receptor-associated factor 3 (TRAF3) and TRAF6 and results in a signal transduction cascade involving activation of transcription factors interferon regulatory factor 3 (IRF3), IRF7, nuclear factor-kappaB (NF-kappaB), and activation protein-1 (AP-1) (1-5). Transcription factors translocate to the nucleus, driving type I interferon (IFN) production, secretion of pro-inflammatory cytokines, and tumor regression (1-5). Furthermore, TRIF can also interact with receptor-interacting serine-threonine kinase 1 (RIP1) and RIP3 leading to reactive oxygen species (ROS) production and apoptosis (1,2,5). Conversely, NF-kappaB transcription can also promote chemokine production and promote the WNT pathway associated with stemness and pro-tumorigenic properties (1,5).

Given the role of TLR3 in immune response, its expression or dysfunction has been associated with a number of pathologies from chronic inflammation to autoimmune disorders and cancer (1-5,7). TLR3 is expressed in many cancer types, often related to viral infection, such as cervical cancer, hepatocellular carcinoma (HCC), melanoma, breast cancer, and prostate cancer (1,5). TLR3 signaling has a dual role in cancer, either contributing to pro- or anti-tumor properties depending on the type of cancer (1,5). Therapeutic targeting the TLR3 signaling pathway is under investigation. TLR3 inhibitors or antagonists are being studied for the treatment autoimmune and inflammatory disorders such as of sepsis and atherosclerosis (2,8). TLR3 agonists, either alone or in combination with immune checkpoint inhibitors or therapeutic agents, are being studied as immunotherapeutic treatments of many cancers such as colorectal cancer, prostate cancer, and melanoma (7).

References

1. Zheng X, Li S, Yang H. Roles of Toll-Like Receptor 3 in Human Tumors. Front Immunol. 2021;12:667454. https://doi.org/10.3389/fimmu.2021.667454

2. Zhuang C, Chen R, Zheng Z, Lu J, Hong C. Toll-Like Receptor 3 in Cardiovascular Diseases. Heart Lung Circ. 2022;S1443-9506(22)00080-4. https://doi.org/10.1016/j.hlc.2022.02.012

3. Bianchi F, Pretto S, Tagliabue E, Balsari A, Sfondrini L. Exploiting poly(I:C) to induce cancer cell apoptosis. Cancer Biol Ther. 2017;18(10):747-756. https://doi.org/10.1080/15384047.2017.1373220

4. Matsumoto M, Seya T. TLR3: interferon induction by double-stranded RNA including poly(I:C). Adv Drug Deliv Rev. 2008;60(7):805-812. https://doi.org/10.1016/j.addr.2007.11.005

5. Muresan XM, Bouchal J, Culig Z, Soucek K. Toll-Like Receptor 3 in Solid Cancer and Therapy Resistance. Cancers (Basel). 2020;12(11):3227. https://doi.org/10.3390/cancers12113227

6. Uniprot (O15455)

7. Le Naour J, Galluzzi L, Zitvogel L, Kroemer G, Vacchelli E. Trial watch: TLR3 agonists in cancer therapy. Oncoimmunology. 2020;9(1):1771143. https://doi.org/10.1080/2162402X.2020.1771143

8. Gao W, Xiong Y, Li Q, Yang H. Inhibition of Toll-Like Receptor Signaling as a Promising Therapy for Inflammatory Diseases: A Journey from Molecular to Nano Therapeutics. Front Physiol. 2017;8:508. https://doi.org/10.3389/fphys.2017.00508

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Reporter Cell Lines are guaranteed for 1 year from date of receipt.

Publications for TLR3 Reporter Cell Line (NBP2-26275)(2)

Publications using NBP2-26275

• Zurcher C, Sauter KS, Schweizer M. Pestiviral Erns blocks TLR-3-dependent IFN synthesis by LL37 complexed RNA. Veterinary Microbiology. 2014-10-13 [PMID: 25457366]
• Chan M, Hayashi T, Mathewson RD et al. Identification of substituted pyrimido[5,4-b]indoles as selective Toll-like receptor 4 ligands. J Med Chem. 2013-06-13 [PMID: 23656327]

FAQs for TLR3 Reporter Cell Line (NBP2-26275). (Showing 1 - 1 of 1 FAQs).

1. Which promoter drives to express TLR on TLR expression vector into the TLR/NF-kB/SEAP Stable Reporter Cell Line. What is the vector which is delivered into this cell line?
   • The TLR vectors are derived from a CMV promoter-based mammalian expression vector. Hence the promoter for the TLR genes is CMV. However, the exact sequence of the vector is proprietary.

Bioinformatics

• Gene Symbol: TLR3