The IFIH1 Knockout HT29 Polyclonal Cells consist of a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colorectal adenocarcinoma cell line, designed to disrupt the IFIH1 gene (encoding MDA5). This polyclonal knockout model provides a heterogeneous loss-of-function system for investigating the roles of MDA5 in antiviral innate immunity and inflammatory signaling within an intestinal epithelial context.
HT29 cells are epithelial and adherent, originally isolated from a primary colon adenocarcinoma of a 44-year-old female. They are widely utilized as a model of intestinal epithelial differentiation, barrier function, and colorectal cancer biology. Their capacity to polarize and form tight junctions makes them suitable for studying immune responses at mucosal surfaces.
IFIH1 encodes MDA5, a cytoplasmic pattern recognition receptor that specifically detects long double-stranded RNA (dsRNA) generated during viral replication. Upon ligand binding, MDA5 oligomerizes and interacts with the mitochondrial adaptor MAVS, which subsequently activates downstream kinases including TBK1 and IKK??. These kinases phosphorylate the transcription factors IRF3 and IRF7, leading to their dimerization and nuclear translocation, where they drive the expression of type I interferons such as IFN-?? and interferon-stimulated genes (e.g., MX1, OAS1). Concurrently, MAVS signaling engages the NF-??B pathway, promoting the production of pro-inflammatory cytokines like IL-6 and TNF-??. Upstream regulators of this cascade include viral dsRNA, type I interferon feedback, and cofactors such as TRIM25 and ZCCHC3, while interacting partners like PACT, LGP2, and DAK modulate MDA5 activity.
In the intestinal epithelial setting, MDA5-dependent signaling is crucial for mounting antiviral defenses and may influence epithelial homeostasis and inflammation. Genetic alterations or dysregulated MDA5 activity are associated with autoimmune conditions such as type I diabetes, systemic lupus erythematosus, Aicardi-Gouti??res syndrome, and Singleton-Merten syndrome. Therefore, disrupting IFIH1 in HT29 cells provides a relevant model to dissect how loss of MDA5 function impacts innate immune activation, apoptosis, and cytokine responses in colorectal epithelial cells, with implications for inflammatory bowel disease and colorectal cancer immunobiology.
This polyclonal knockout cell population is suited for a range of experimental applications, including transfection with the synthetic dsRNA analog poly(I:C) to assess interferon-stimulated gene induction by RT-qPCR or RNA-seq, western blot analysis of phospho-IRF3, ELISA detection of secreted IFN-??, and infection studies with viruses such as encephalomyocarditis virus (EMCV). The model also supports investigation of oncolytic virus sensitivity, dsRNA-induced apoptosis, and innate immune checkpoint mechanisms in colorectal cancer. For additional information about this product, please contact Ascent Research.
