The ACOX3 Knockout HT29 Polyclonal Cells consist of a polyclonal CRISPR/Cas9-edited HT29 population with targeted disruption of the ACOX3 gene. This knockout model ablates pristanoyl-CoA oxidase activity, enabling investigation of peroxisomal beta-oxidation in a human colorectal adenocarcinoma background. As a polyclonal pool, the cells avoid clonal artifacts and provide a robust system for studying loss-of-function effects on branched-chain fatty acid metabolism and lipid homeostasis.
The HT29 host cell line is derived from a primary human colon adenocarcinoma and retains intestinal epithelial characteristics. It is widely used in colon cancer research, metabolic studies, and differentiation assays. HT29 cells can be maintained as an undifferentiated monolayer or induced to differentiate, offering a flexible platform to examine how ACOX3 knockout influences tumor metabolism and cellular differentiation programs. Their established use in lipid metabolism and peroxisomal function studies streamlines experimental integration.
ACOX3 encodes pristanoyl-CoA oxidase, which catalyzes the peroxisomal desaturation of pristanoyl-CoA to 2,3-pristanoyl-CoA. This enzyme works sequentially with D-bifunctional protein (HSD17B4) and peroxisomal thiolase (ACAA1) to produce acetyl-CoA and propionyl-CoA. ACOX3 expression is regulated by PPAR?? and PPAR?? agonists, as well as by FOXA2 and HNF4??. Its activity depends on peroxisomal biogenesis factors (PEX proteins), linking ACOX3 to overall peroxisomal integrity and cellular lipid homeostasis.
ACOX3 knockout in HT29 cells disrupts peroxisomal branched-chain fatty acid oxidation, leading to pristanic acid accumulation and altered lipid profiles. This model permits the study of peroxisomal dysfunction in colorectal cancer metabolism, including effects on proliferation, differentiation, and stress responses. The HT29 background is particularly relevant for investigating the interplay between peroxisomal activity and oncogenic signaling, helping to identify metabolic vulnerabilities in colorectal tumors.
Key applications include quantitative radiolabeled pristanate oxidation assays to measure peroxisomal beta-oxidation activity, Western blotting for ACOX3 and peroxisomal markers such as PMP70, and immunofluorescence to visualize peroxisome morphology and abundance. The polyclonal knockout population is well-suited for RT-qPCR analysis of PPAR target genes, comprehensive lipidomics profiling to monitor changes in fatty acid species and acylcarnitines, and viability assays under lipid stress to probe metabolic dependencies. This model also serves as a platform for drug screening aimed at restoring or modulating peroxisomal function. For additional information or customization, please contact Ascent Research.
