The ACSL4 Knockout HCT 116 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HCT 116 colorectal carcinoma line, providing a loss-of-function model for acyl-CoA synthetase long-chain family member 4 (ACSL4). This product enables researchers to interrogate ACSL4-dependent mechanisms in lipid metabolism, ferroptosis, and oncogenic signaling without the need for clonal isolation, offering a heterogeneous knockout background that mirrors population-level gene disruption effects. The polyclonal format is particularly suited for pooled functional screens and studies where polygenic variation within the edited population may yield more robust phenotypic assessments.
The host HCT 116 cell line is an epithelial colorectal adenocarcinoma model originating from a human male, characterized by a mutant KRAS (G13D) allele that drives constitutive MAPK pathway activation. This genetic background promotes aggressive tumorigenicity and metabolic reprogramming, making it a well-established platform for investigating colorectal cancer biology. HCT 116 cells exhibit adherent growth, rapid proliferation, and sensitivity to oxidative stress, providing a pertinent context for dissecting the role of ACSL4 in ferroptosis and lipid-mediated cell death.
ACSL4 catalyzes the ligation of long-chain polyunsaturated fatty acids, such as arachidonic acid, to coenzyme A, thereby generating acyl-CoA substrates for phospholipid esterification via lysophosphatidylcholine acyltransferase 3 (LPCAT3). This enzymatic cascade enriches cellular membranes with oxidizable phospholipids, predisposing cells to ferroptosis initiated by lipid peroxidation. ACSL4 expression is transcriptionally regulated by peroxisome proliferator-activated receptors ?? and ?? (PPAR??/??), sterol regulatory element-binding protein 1 (SREBP1), and tumor protein p53 (TP53). Its activity interfaces with LPCAT3, arachidonate lipoxygenases ALOX5 and ALOX15, and glutathione peroxidase 4 (GPX4), which counteracts lipid hydroperoxide accumulation. The pathway is further integrated with system Xc?-mediated cystine import, highlighting a regulatory node where ACSL4, LPCAT3, ALOX15, GPX4, and lipid hydroperoxides coordinate ferroptotic sensitivity.
Within the colorectal cancer context, ACSL4-mediated lipid remodeling influences ferroptosis susceptibility, eicosanoid synthesis, and phospholipid diversity, all of which impact tumor cell survival under metabolic stress. The HCT 116 KRAS mutation drives aberrant lipid uptake and utilization, potentially intersecting with ACSL4 function to modulate therapy response. Knocking out ACSL4 in these cells allows dissection of how oncogenic signaling??via SREBP1 or TP53??alters lipid metabolic reprogramming and ferroptosis execution, offering insight into vulnerabilities that may be exploited for therapeutic intervention.
This polyclonal knockout cell population is optimized for diverse ferroptosis mechanistic studies, including lipid peroxidation detection with BODIPY 581/591 C11, cell viability assays using erastin or RSL3, and western blotting for GPX4 expression. Researchers can employ RT-qPCR to verify ACSL4 transcript disruption, and complementary techniques such as migration/invasion assays, colony formation, flow cytometry for cell death, and LC-MS-based phospholipid profiling. The model also supports drug resistance research and ferroptosis modulator screening. For further details, please contact Ascent Research.
