The ACAA2 Knockout HAP1 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal population carrying targeted disruption of the ACAA2 gene in the human near-haploid HAP1 cell line. This loss-of-function model is generated without single-cell cloning, providing a heterogeneous mixture of knockout alleles useful for population-based functional assays. The polyclonal format preserves genetic diversity while enabling robust interrogation of ACAA2-dependent pathways, particularly those involved in mitochondrial fatty acid metabolism. This product is suitable for applications in metabolic research, cancer biology, and drug discovery, where acute ablation of ACAA2 can reveal its role in lipid catabolism and energy homeostasis.
The host HAP1 cell line is a near-haploid chronic myeloid leukemia (CML) model derived from the KBM-7 line, characterized by BCR-ABL positivity and p53 deficiency. Its near-haploid karyotype simplifies genetic perturbation analysis, as most genes are present in a single copy, reducing the complexity associated with diploid genomes. HAP1 cells are widely employed in functional genomics screens, drug sensitivity profiling, and haploid genetic screens, making them an ideal background for dissecting metabolic vulnerabilities. The mesenchymal-like morphology and adherent growth facilitate a variety of downstream assays, including live-cell metabolic analysis.
ACAA2 encodes mitochondrial 3-ketoacyl-CoA thiolase, an enzyme that catalyzes the final step of each cycle of fatty acid ??-oxidation. It cleaves 3-ketoacyl-CoA substrates to produce acetyl-CoA and a shortened acyl-CoA, thereby feeding carbon units into the TCA cycle and ketone body synthesis. ACAA2 is transcriptionally regulated by PPAR?? and its coactivator PGC-1??, activated by AMPK-mediated energy sensing, and modulated by hormonal signals such as insulin and glucagon. Within the mitochondrial trifunctional protein complex, ACAA2 physically interacts with HADHA, HADHB, ECHS1, and HSD17B10 to coordinate efficient ??-oxidation. Disruption of ACAA2 disrupts this multienzyme machinery, leading to accumulation of upstream intermediates and reduced acetyl-CoA output, impairing cellular energy production and lipid-dependent metabolic flexibility.
In the HAP1 background, which retains oncogenic BCR-ABL signaling and lacks p53-mediated metabolic checkpoints, ACAA2 knockout provides a powerful tool to study how cancer cells rewire lipid utilization. The near-haploid nature ensures that a single disruptive editing event suffices to ablate gene function, enabling clear phenotypic readouts in metabolic assays. This model is particularly relevant for investigating the intersection of fatty acid oxidation with leukemogenic signaling, as BCR-ABL-driven cells often exhibit altered lipid metabolism. Researchers can use these polyclonal knockout cells to explore compensatory pathways, such as ketogenesis or anaplerotic reactions, that maintain ATP and biosynthetic precursor pools when ??-oxidation is compromised.
Typical applications include fatty acid oxidation flux assays using radiolabeled or stable isotope-labeled fatty acids, Seahorse respirometry to measure mitochondrial oxygen consumption, and targeted metabolomics or lipidomics to profile acyl-CoA species and downstream metabolites. Furthermore, these cells are amenable to high-content screening for metabolic disease drug candidates, viability assays (e.g., MTT), and integration with CRISPR-based synthetic lethality screens. Validation of knockout efficiency can be performed via next-generation sequencing and western blotting, while RT-qPCR can assess transcriptional adaptations. By providing a genetically defined, polyclonal HAP1 knockout model, this product accelerates discovery in metabolic disorders and cancer metabolism. For additional technical information or ordering inquiries, please contact Ascent Research.
