The CA2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population designed to achieve target-gene disruption of the CA2 locus in the HAP1 human near-haploid cell line. This loss-of-function model abolishes expression of carbonic anhydrase II, enabling researchers to investigate the enzyme’s roles without residual wild-type activity. The polyclonal format provides a population-level knockout, suitable for pooled functional assays, where consistent genetic ablation is maintained across the culture.
HAP1 cells are derived from KBM-7 chronic myeloid leukemia cells and possess a near-haploid karyotype, which simplifies genetic manipulation by eliminating the need for biallelic editing. This haploid background ensures a clean knockout phenotype, as there is no second allele to compensate. Their robust growth characteristics and amenability to CRISPR/Cas9 editing have established HAP1 as a powerful platform for genetic screens and mechanistic studies, particularly in pathways where complete loss-of-function is critical for phenotypic analysis.
CA2 encodes carbonic anhydrase II, a cytosolic enzyme that catalyzes the reversible hydration of CO2 to HCO3- and H+, playing a central role in pH homeostasis, respiration, ion transport, and bone resorption. The enzyme functions within a bicarbonate transport metabolon by directly interacting with the anion exchangers and cotransporters such as SLC4A1 (AE1) and SLC4A4 (NBCe1), efficiently channeling bicarbonate across membranes. Its activity is regulated by upstream factors including 1,25-dihydroxyvitamin D3, parathyroid hormone, epidermal growth factor, and physiological parameters like CO2 concentration and pH. Downstream, CA2-mediated proton generation drives vacuolar H+-ATPase activity in osteoclasts and supports renal bicarbonate reclamation, highlighting its integration into systemic acid-base balance.
In the HAP1 context, knockout of CA2 creates a uniquely defined cellular model since the haploid genome yields a complete loss-of-function without confounding wild-type expression. This system is particularly valuable for dissecting carbonic anhydrase II deficiency-related pathologies, including osteopetrosis autosomal recessive type 3, renal tubular acidosis, and cerebral calcification. The model also enables precise investigation of bicarbonate transport mechanisms and the evaluation of carbonic anhydrase inhibitors such as acetazolamide, as off-target effects are minimized in a defined genetic background.
This CA2 knockout cell pool supports diverse research applications, including functional genomics screens, drug discovery for carbonic anhydrase modulators, and studies of cancer cell metabolism. Representative assays include intracellular pH measurements using pH-sensitive dyes, spectrophotometric carbonic anhydrase activity assays, bicarbonate transport kinetics, RT-qPCR and Western blotting for pathway validation, and proliferation or viability tests under metabolic stress. Researchers can employ this model to elucidate pH regulation in leukemic cells, screen for novel inhibitors, or study the interplay between carbonic anhydrase II and interacting partners such as aquaporin-1 and SLC26A7. For additional information, please contact Ascent Research.
