The HACL2 Knouckout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population in which the HACL2 gene has been disrupted in the HT29 colorectal adenocarcinoma cell line. This product provides a heterogeneous mixture of edited alleles that collectively ablate HACL2 protein expression, preserving population-level variability while eliminating target-gene function. The CRISPR/Cas9-mediated gene disruption ensures efficient loss of function without introducing exogenous transgenes, making the model suitable for studies requiring native cellular context.
HT29 cells, originally derived from a 44-year-old Caucasian female with colorectal adenocarcinoma (ATCC HTB-38), are a widely used intestinal epithelial model. These cells retain the capacity to undergo enterocytic differentiation and form polarized monolayers, reflecting key features of the colonic epithelium. Their robust growth and compatibility with genome editing make them a valuable platform for investigating cancer cell biology and metabolic pathways.
HACL2 encodes 2-hydroxyacyl-CoA lyase, a peroxisomal enzyme that catalyzes the cleavage of 2-hydroxyacyl-CoAs during alpha-oxidation of phytanic acid. This reaction produces formyl-CoA and long-chain aldehydes, which feed into further oxidative metabolism. HACL2 activity is regulated upstream by PPAR??, a nuclear receptor activated by phytanic acid, and depends on peroxisomal import receptors PEX5 and PEX7. Interacting factors include phytanoyl-CoA hydroxylase (PHYH/PAHX) and the homologous lyase HACL1. Knockout of HACL2 results in accumulation of 2-hydroxyacyl-CoA intermediates, impairing branched-chain fatty acid degradation and potentially disrupting cellular energy metabolism and redox balance.
In HT29 cells, HACL2 knockout poses a specific disruption to peroxisomal alpha-oxidation, which is particularly relevant for modeling phytanic acid storage disorders such as adult Refsum disease. Given that colorectal cancer cells exhibit altered lipid metabolism and peroxisomal function, this polyclonal knockout model enables the study of how loss of this lyase influences tumor cell metabolic rewiring, sensitivity to oxidative stress, and peroxisome?Cmitochondria crosstalk. The model thus bridges inherited metabolic disease mechanisms and cancer biology.
Typical applications include phytanic acid treatment and viability assays to assess metabolic vulnerability, LC?CMS-based fatty acid profiling to monitor 2-hydroxyacyl-CoA accumulation, and Seahorse metabolic flux analysis to evaluate changes in mitochondrial respiration and glycolysis. Knockout efficiency can be validated by RT-qPCR and western blotting, while immunofluorescence for peroxisomal markers (e.g., PEX14) enables morphological assessment. These cells are also suited for drug discovery targeting peroxisomal disorders and biomarker identification. For further technical details, please contact Ascent Research.
