CCS Knockout HAP1 Polyclonal Cells provide a CRISPR/Cas9-edited polyclonal knockout population targeting the CCS gene in human HAP1 cells. This loss-of-function model enables investigation of copper chaperone function and oxidative stress regulation. The polyclonal format offers a heterogeneous pool of edited cells, supporting robust experimental reproducibility without clonal selection bias.
HAP1 cells are a near-haploid human cell line derived from KBM-7 chronic myeloid leukemia cells. Their near-haploidy simplifies gene targeting and phenotypic analysis, as genes are usually present in single copies. Retaining hematopoietic signaling features, HAP1 cells are extensively used in genetic screens, drug discovery, and functional genomics, offering a tractable platform for studying gene function in hematopoietic contexts.
CCS is a copper chaperone that specifically delivers copper to Cu/Zn superoxide dismutase (SOD1), enabling its enzymatic dismutation of superoxide radicals (O2??) into hydrogen peroxide (H2O2) and oxygen. CCS expression is regulated by metal-responsive transcription factor-1 (MTF1), nuclear factor erythroid 2-related factor 2 (NRF2), hypoxia, and copper levels. Through direct interaction with SOD1, copper ions, and heat shock protein 70 (HSP70), CCS facilitates copper transfer, which is essential for SOD1 activation. Active SOD1 converts superoxide to H2O2, subsequently detoxified by catalase and glutathione peroxidase. Disruption of CCS prevents copper incorporation into SOD1, leading to superoxide accumulation, elevated reactive oxygen species (ROS), and oxidative damage, thereby impacting cell survival and stress responses.
In the HAP1 near-haploid background, CCS knockout provides a clean model for dissecting copper-dependent SOD1 activation in hematopoietic cells. This system is relevant for studying oxidative stress regulation in blood lineages and neurodegeneration models such as amyotrophic lateral sclerosis (ALS) with SOD1 mutations. Loss of CCS abolishes SOD1 activity, enabling examination of superoxide-mediated signaling, oxidative DNA damage, and compensatory antioxidant pathways. The polyclonal nature reflects heterogeneous editing, offering a realistic model for pharmacological screening.
Applications include Western blotting for CCS and SOD1, SOD1 enzymatic activity assays, and superoxide detection via dihydroethidium. Cellular ROS measurements, oxidative stress markers (8-oxoguanine, protein carbonyls), and cell viability under oxidative challenge (e.g., paraquat, H2O2) are readily performed. This model also supports copper content analysis and co-immunoprecipitation of CCS-SOD1 complexes. Key research areas encompass oxidative stress mechanisms, SOD1-linked neurodegeneration, copper metabolism, antioxidant development, and cancer oxidative vulnerability. For more information, contact Ascent Research.
