HEK-Blue™ KO-ALPK1 Cells
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Cat.code:
hkb-koalpk
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ABOUT
Knockout of ALPK1 or TIFA in HEK-293 cells
The ALPK1-TIFA signaling axis is an important cytoplasmic surveillance pathway of pathogenic Gram-negative bacteria. To foster research on this pathway, InvivoGen provides cellular tools that have been generated from the parental HEK-Blue™ Null1-v Cells, a human embryonic kidney (HEK)-293 derived cell line, through the stable knockout (KO) of the ALPK1 or TIFA gene. These cell lines express a secreted embryonic alkaline phosphatase (SEAP) under the control of an NF-κB-inducible promoter. Levels of SEAP in the supernatant can be easily determined with HEK-Blue™ Detection, cell culture medium for real-time detection, or QUANTI-Blue™ detection reagent (see figures).
• HEK-Blue™ KO-ALPK1 Cells - Knockout (KO) of the ALPK1 gene
• HEK-Blue™ KO-TIFA Cells – Knockout (KO) of the TIFA gene
Unlike their parental cell line HEK-Blue™ Null1-v, both HEK Blue™ KO-ALPK1 and HEK Blue™ KO-TIFA cells do not respond to cytosolic ADP-Heptose. However, they do respond to other NF‑κB inducing cytokines such as TNF-α and IL-1β.
The ALPK1-TIFA signaling axis
Alpha protein Kinase 1 (ALPK1) and TRAF interacting forkhead-associated protein A (TIFA) form an important cytoplasmic surveillance pathway of pathogenic Gram-negative bacteria such as Shigella flexneri and Helicobacter pylori [1,2].
ALPK1 is a critical cytoplasmic alpha kinase responsible for the recognition and binding of bacterial sugar metabolites. ALPK1 recognizes and binds directly to the metabolic intermediates in lipopolysaccharide (LPS) biosynthesis, the potent ADP-β-d-manno-heptose (ADP-Heptose) and the enzymatically converted d-glycero-β-d-manno-heptose-1,7-bisphosphate (HBP) [3]. ADP-Heptose and HBP can be delivered to the host cell cytoplasm in a number of different ways including type III and IV secretion systems (e.g. Yersinia pseudotuberculosis and H. pylori, respectively), as well as by endocytosed bacteria [2-3]. Notably, ADP-Heptose can freely penetrate the cell membrane, whereas HBP requires the use of pore-forming agents [3, 4].
Activated ALPK1 phosphorylates TRAF interacting forkhead-associated protein A (TIFA), inducing its oligomerization and the formation of 'TIFAsomes', which in turn recruits and activates TRAF6. Ultimately, the ALPK1-TIFA signaling axis results in the initiation of an NF-κB dependent pro-inflammatory response [1-4].
Targeting this pathway will be important in the development of novel treatments for modulating inflammation caused by bacterial infection as well as diseases associated with its dysregulation such as gastric cancer.
Features of HEK-Blue™ KO-ALPK1 and KO-TIFA Cells:
- Verified KO of the ALPK1 or TIFA gene (DNA sequencing, PCR, and functional assays)
- Complete abrogation of the response to the LPS-intermediary metabolite ADP-Heptose
- Readily assessable NF-κB-dependent SEAP reporter activity
Features of HEK-Blue™ Null1-v Cells:
- Fully functional ALPK1-TIFA signaling pathway
- Responds to the LPS-intermediary metabolite ADP-Heptose
- Readily assessable NF-κB-dependent SEAP reporter activity
References:
1. Garcia-Weber, D. et al. 2018. ADP-heptose is a newly identified pathogen-associated molecular pattern of Shigella flexneri. EMBO Rep 19
2. Zimmermann, S. et al. 2017. ALPK1- and TIFA-Dependent Innate Immune Response Triggered by the Helicobacter pylori Type IV Secretion System. Cell Rep 20, 2384-2395.
3. Zhou, P. et al. 2018. Alpha-kinase 1 is a cytosolic innate immune receptor for bacterial ADP-heptose. Nature 561, 122-126.
4. Pfannkuch, L. et al. 2019. ADP heptose, a novel pathogen-associated molecular pattern identified in Helicobacter pylori. FASEB J, fj201802555R.
Disclaimer: These cells are for internal research use only and are covered by a Limited Use License (See Terms and Conditions). Additional rights may be available.
SPECIFICATIONS
Specifications
DNA sequencing, PCR, functional assays
Complete DMEM (see TDS)
Verified using Plasmotest
Each lot is functionally tested and validated.
CONTENTS
Contents
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Product:HEK-Blue™ KO-ALPK1 Cells
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Cat code:hkb-koalpk
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Quantity:3-7 x 10^6 cells
- 1 ml of Zeocin® (100 mg/ml)
- 1 ml of Normocin™ (50 mg/ml). Normocin™ is a formulation of three antibiotics active against mycoplasmas, bacteria, and fungi.
- 1 pouch of HEK-Blue™ Detection (cell culture medium for real-time detection of SEAP)
Shipping & Storage
- Shipping method: Dry ice
- Liquid nitrogen vapor
- Upon receipt, store immediately in liquid nitrogen vapor. Do not store cell vials at -80°C.
Storage:
Caution:
DOCUMENTS
Documents
Technical Data Sheet
Validation Data Sheet
Safety Data Sheet
Certificate of analysis
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