The ACO1 Knockout HT29 Polyclonal Cells are a polyclonal population of HT29 human colorectal adenocarcinoma cells engineered with CRISPR/Cas9-mediated disruption of the ACO1 gene. This knockout ablates both cytosolic aconitase activity and iron regulatory protein 1 (IRP1) function, enabling dissection of iron-dependent pathways and TCA cycle metabolism in a colorectal cancer context.
HT29 is a widely used colorectal adenocarcinoma cell line that is epithelial in origin, microsatellite stable, and harbors oncogenic mutations in BRAF (V600E), APC, and TP53. These cells serve as a model for intestinal epithelial differentiation and colorectal carcinogenesis, and are frequently employed to study signal transduction, drug responses, and metabolic reprogramming.
The ACO1 protein acts as a cytosolic aconitase in the TCA cycle when iron is sufficient and an iron-sulfur (Fe-S) cluster is intact. Under low iron conditions, cluster disassembly converts the protein to IRP1, which binds to iron-responsive elements (IREs) in mRNAs encoding proteins such as TFRC (transferrin receptor), ferritin light and heavy chains (FTL, FTH1), ferroportin (SLC40A1), and DMT1 (SLC11A2). IRP1 activity is regulated by intracellular iron levels, hypoxia (via HIF1A), nitric oxide, and oxidative stress, and it cooperates with IRP2 (IREB2) and the Fe-S cluster assembly machinery. Loss of ACO1 therefore disrupts the post-transcriptional coordination of iron uptake, storage, and efflux and may redirect TCA cycle flux.
In the HT29 colorectal cancer background, ACO1 knockout is particularly relevant because iron metabolism is frequently dysregulated to support proliferation and survival. HT29 cells with BRAF V600E and mutant p53 may exhibit altered dependency on iron homeostasis and redox control, making this model valuable for investigating how IRP1 loss affects colorectal cancer cell growth, differentiation, and response to iron-targeted interventions. The dual metabolic and regulatory functions of ACO1 also permit examination of the interplay between iron handling and central carbon metabolism in a genetically defined adenocarcinoma line.
This polyclonal knockout cell population is suitable for a variety of functional studies, including Western blotting for ACO1, ferritin, and TFRC; aconitase enzyme activity assays; iron uptake measurements with 55Fe; RT?qPCR of IRE-containing transcripts; and flow cytometric assessment of surface TFRC. Applications range from exploring iron metabolism in colorectal cancer and intestinal epithelial iron homeostasis to screening for modulators of iron regulatory proteins and analyzing TCA cycle rewiring under iron perturbation. For further information, please contact Ascent Research.
