ACO2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population generated in the HAP1 human cell line. These cells harbor a targeted disruption of the ACO2 gene, resulting in a loss-of-function model for studying mitochondrial aconitase and iron regulatory protein 1 functions. The polyclonal format provides a heterogeneous pool of edited cells, ideal for pooled genetic screens and functional genomics studies.
The HAP1 cell line is a near-haploid human cell line derived from the male chronic myeloid leukemia cell line KBM-7. Its haploid nature makes it a powerful model for genetic screening and knockout studies, as it allows efficient CRISPR-mediated gene disruption due to the presence of a single set of chromosomes, reducing redundancy. HAP1 cells are widely used in functional genomics, drug target validation, and pathway analysis.
ACO2 encodes mitochondrial aconitase, which catalyzes the reversible isomerization of citrate to isocitrate in the TCA cycle, a central metabolic pathway. Additionally, ACO2 functions as a cytosolic iron-regulatory protein 1 (IRP1) that binds to iron-responsive elements (IREs) in mRNAs, post-transcriptionally regulating iron homeostasis genes such as ferritin, transferrin receptor, and ferroportin. ACO2 is regulated by iron availability and oxidative stress, and its activity is influenced by upstream factors including NRF1 and NF-??B. Downstream, ACO2-derived isocitrate feeds into the TCA cycle for ??-ketoglutarate production, linking to oxidative phosphorylation and mitochondrial metabolism. ACO2 interacts with citrate, iron-sulfur clusters, and other TCA enzymes like IDH1 and IDH2, and its IRP1 form cooperates with IRP2 to modulate cellular iron levels.
In the HAP1 background, ACO2 disruption allows dissection of its dual role in metabolism and iron regulation. The near-haploid context ensures that each cell carries a single engineered ACO2 allele, facilitating clear genotype-phenotype correlations. This model is particularly valuable for studying diseases linked to ACO2 dysfunction, such as infantile cerebellar-retinal degeneration and neurodegenerative disorders, as well as cancer metabolic reprogramming, where aconitase activity and iron metabolism are often altered. The polyclonal nature enables pooled screening approaches to identify genetic interactions and modulators of ACO2-dependent pathways.
Researchers can employ these cells in aconitase enzyme activity assays, Seahorse metabolic flux analysis, and iron uptake/efflux assays to directly assess ACO2 function. Western blotting, RT-qPCR, and immunofluorescence can validate knockout and downstream effects on target genes like ferritin and transferrin receptor. RNA electrophoretic mobility shift assays (EMSA) are suitable for studying IRP1-IRE binding. The cells are also amenable to genetic interaction screens and cell viability assays under metabolic stress. For further details or to enquire about this product, please contact Ascent Research.
