The HACL1 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout population generated from the HT29 human colorectal adenocarcinoma cell line, with targeted disruption of the HACL1 gene encoding 2-hydroxyacyl-CoA lyase. This heterogeneous cell pool provides a physiologically relevant model for studying peroxisomal alpha-oxidation deficiency in an intestinal epithelial background, circumventing clonal selection while maintaining population-level loss of function.
HT29 is an adherent epithelial cell line originally derived from a primary colorectal tumor. It is widely employed as a model for intestinal epithelial biology and colorectal cancer research due to its capacity to form mucin-secreting monolayers, express enterocytic markers, and recapitulate aspects of colon carcinoma. Furthermore, HT29 cells contain a subpopulation that can undergo enterocytic differentiation, providing an opportunity to study metabolic alterations in a more polarized intestinal state. HT29 cells are valuable for investigating cellular processes such as nutrient transport, barrier function, and lipid metabolism within the intestinal epithelium.
The HACL1 enzyme catalyzes thiamine pyrophosphate-dependent cleavage of 2-hydroxyacyl-CoA to fatty aldehyde and formyl-CoA, a critical step in peroxisomal alpha-oxidation of phytanic acid. PHYH hydroxylates phytanoyl-CoA upstream, a process requiring PEX7, while ALDH3A2 oxidizes the fatty aldehyde product. HACL1 transcription is regulated by PPAR??, which is activated by phytanic acid and fibrates, placing it within a key lipid metabolism network.
Disruption of HACL1 in HT29 polyclonal cells halts alpha-oxidation, leading to accumulation of phytanic acid and 2-hydroxyphytanoyl-CoA, recapitulating the Refsum disease metabolic defect. This model allows investigation of how peroxisomal dysfunction impacts colorectal cancer cell biology, including effects on lipid homeostasis and PPAR??-dependent signaling. Prolonged phytanic acid accumulation may activate PPAR?? and induce oxidative stress, enabling studies of lipid-induced cellular damage in intestinal cells.
This polyclonal knockout is suitable for Refsum disease modeling, phytanic acid toxicity studies, drug screening for peroxisomal disorders, and research on peroxisomal roles in colorectal cancer. Verification assays include Western blot, RT-qPCR, HACL1 enzyme activity, GC-MS for phytanic acid, immunofluorescence for PMP70, viability under phytanic acid stress, and PPAR?? luciferase reporter assays. For further information, contact Ascent Research.
