The CALCOCO2 Knockout HAP1 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout population of the human near-haploid HAP1 cell line, designed for targeted disruption of the CALCOCO2 gene encoding the autophagy receptor NDP52. This polyclonal knockout cell model provides a genetically heterogeneous loss-of-function population suitable for studying gene function without clonal selection, enabling robust and reproducible assessments of CALCOCO2-dependent biological processes in a myeloid leukemia background.
The HAP1 host cell line is a near-haploid human myeloid leukemia cell line derived from the KBM-7 chronic myeloid leukemia cells. Its near-haploid karyotype simplifies genetic manipulation and functional genomics studies, as recessive mutations are directly phenotypically exposed. HAP1 cells are widely employed in leukemia research, drug sensitivity profiling, and high-throughput genetic screens, offering a physiologically relevant yet experimentally tractable model for dissecting signaling networks and cancer biology.
CALCOCO2 encodes NDP52, a key selective autophagy receptor that recognizes ubiquitinated intracellular pathogens and protein aggregates, facilitating their degradation by recruiting the autophagic machinery through LC3-interacting region (LIR) motifs. NDP52 directly interacts with members of the LC3/GABARAP family (MAP1LC3A, MAP1LC3B, GABARAP) and the autophagy adaptor p62/SQSTM1 to promote autophagosome formation. NDP52 functions as a scaffold for TBK1 kinase and interacts with NAP1 and SINTBAD to modulate NF-??B signaling. Upstream regulators such as NF-??B, TBK1, interferon-gamma (IFNG), and lipopolysaccharide (LPS) control NDP52 expression and activation, while downstream, NDP52 influences LC3 lipidation, degradation of ubiquitinated substrates, and NF-??B transcriptional activity via the IKK complex and p65.
In the HAP1 near-haploid background, disruption of CALCOCO2 provides a powerful system to dissect the interplay between selective autophagy and innate immune signaling in leukemia cells. The knockout model permits unambiguous assessment of NDP52-dependent clearance of intracellular bacteria (e.g., Salmonella) and protein aggregates, as well as its contribution to NF-??B activation and inflammatory gene expression. Given the implication of NDP52 in Crohn??s disease, infectious diseases, and cancer, this knockout cell population enables mechanistic studies linking autophagy defects to disease pathogenesis in a human leukemic context.
Typical research applications include western blotting for LC3 lipidation and p62 degradation, co-immunoprecipitation to probe ubiquitin binding and TBK1 interaction, and immunofluorescence microscopy for autophagosome formation. Further assays include RT-qPCR for NF-??B target genes, flow cytometry for viability and apoptosis, bacterial invasion assays, and drug sensitivity tests with autophagy modulators. This polyclonal knockout population is a versatile tool for drug discovery and functional genomics. For further details, contact Ascent Research.
