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NF-κB–SEAP and IL-8–Lucia reporter HEK293 cells expressing murine TLR7

Signaling pathways in HEK-Blue-Lucia™ mTLR7 cells

HEK-Blue-Lucia™ mTLR7 cells were engineered from the human embryonic kidney HEK293 cell line to study the mouse Toll-like receptor 7 (TLR7). This important pattern recognition receptor (PRR) recognizes viral single-stranded RNA (ssRNA) structures and subsequently triggers NF-κB and IRF immune responses [1].

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Description

HEK-Blue-Lucia™ mTLR7 cells were generated from the HEK-Blue-Lucia™ Null cell line harboring two inducible reporter genes. This feature allows the double readout of the NF-κB/AP-1 pathway, by monitoring the SEAP (secreted embryonic alkaline phosphatase) or Lucia luciferase activities. HEK-Blue-Lucia™ mTLR7 cells also stably express the mTLR7 gene. Due to a triple knockout (KO) of TLR3, TLR5, and TNFR, this cell line allows for the independent study of TLR7.

Stimulation of HEK-Blue-Lucia™ mTLR7 cells with TLR7 agonists (e.g. R848) triggers the activation of the artificial NF-κB-inducible promoter and the subsequent production of SEAP. It also promotes the expression of Lucia luciferase, which is knocked in (KI) downstream of the endogenous (more physiological) IL-8 promoter (see figures).

IL-8 (interleukin 8) is a chemokine produced in response to TLR agonists in an NF-κB/AP-1-dependent manner [2-3]. This feature enables the double readout study of the NF-κB/AP-1 pathway, by monitoring the activity of SEAP and Lucia luciferase using QUANTI-Blue™ Solution (SEAP detection reagent) or QUANTI-Luc™ 4 Lucia/Gaussia (luciferase detection reagent), respectively. Thus, you may choose the readout depending on your laboratory equipment utilizing a spectrophotometer for SEAP or a luminometer for Lucia luciferase detection.

Read our review on TLR7 & TLR8

Key features

Stable overexpression of mTLR7
Verified KO for the TLR3, TLR5, and TNFR genes
Functionally validated using a selection of PRR ligands and cytokines
Readily assessable NF-κB activation by assessing the SEAP and/or Lucia luciferase activities

Applications

Defining the role of TLR7-dependent NF-κB signaling pathway
Screening for novel TLR7 agonists and inhibitors
Choice of readout depends on the laboratory equipment (spectrophotometer for SEAP or luminometer for Lucia luciferase detection).

* Note: This cell line has been renamed. It was formerly known as "HEK-Dual™ mTLR7 (NF/IL8)". The cat. code remains (hkd-mtlr7ni) unchanged.

References:

1. Georg P. & Sander L.E., 2019. Innate sensors that regulate vaccine responses. Curr. Op. Immunol. 59:31.
2. Roebuck KA. 1999. Regulation of interleukin-8 gene expression. J Interferon Cytokine Res:429-38.
3. Ohta K, et al. 2014. Toll-like receptor (TLR) expression and TLR‑mediated interleukin-8 production by human submandibular gland epithelial cells. Mol Med Rep. (5):2377-82.

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