This CRISPR/Cas9-edited polyclonal knockout cell population is derived from the HAP1 human cell line and designed to disrupt the ATG7 gene. The pooled format provides a heterogeneous collection of cells carrying knockout-inducing edits at the ATG7 locus, enabling robust loss-of-function studies without single-cell cloning. This model is ideally suited for investigating the consequences of ATG7 deficiency in a near-haploid genetic context, permitting straightforward genotype?Cphenotype correlation in autophagy research and beyond.
HAP1 cells are a near-haploid human cell line originally derived from the KBM-7 chronic myeloid leukemia lineage. Characterized by a single copy of most chromosomes, the line is exceptionally amenable to CRISPR/Cas9-mediated gene disruption: a single successful editing event can yield functional knockout. This genetic simplicity minimizes confounding effects from heterozygous alleles, making HAP1 a preferred host for functional genomics, drug target validation, and pathway dissection where unambiguous loss-of-function states are required.
ATG7 acts as an essential E1-like enzyme that activates two ubiquitin-like conjugation cascades central to autophagosome formation. It catalyzes ATP-dependent activation of ATG12, which is transferred to ATG10 and ultimately conjugated to ATG5, forming the ATG12?CATG5-ATG16L1 complex. Concurrently, ATG7 activates LC3 family members for transfer to ATG3 and subsequent lipidation to phosphatidylethanolamine, generating the lipidated LC3-II form associated with autophagic membranes. Upstream, ULK1 complex and AMPK stimulate ATG7 activity under starvation or growth factor deprivation, whereas mTORC1 exerts inhibitory control. Downstream, functional ATG7 is required for recruitment of selective autophagy receptors such as p62/SQSTM1 and for the lipidation of LC3 that marks autophagosome structures.
Disruption of ATG7 in the HAP1 background abrogates autophagic flux, leading to accumulation of cargo receptors like p62, failure to generate LC3-II, and impaired clearance of protein aggregates and damaged organelles. The near-haploid karyotype ensures that loss-of-function edits within the polyclonal pool are expressed phenotypically without interference from a wild-type allele. Consequently, this model provides a clean system to study autophagy-dependent processes in both basal and stress conditions, including nutrient deprivation and proteotoxic stress, and serves as a reliable control for ATG7-dependent pathway experiments.
This ATG7 knockout HAP1 polyclonal cell pool supports a broad range of applications: fundamental autophagy research, cancer biology studies, neurodegeneration modeling, drug screening for autophagy modulators, and functional analysis of ATG7 in infection and immunity. Representative assays include Western blotting for LC3-II and p62, fluorescence microscopy of GFP-LC3 puncta, autophagic flux measurements with bafilomycin A1, cell viability under starvation, co-immunoprecipitation of ATG7 interactors, and RT-qPCR profiling of autophagy-related genes. For detailed specifications and purchasing, contact Ascent Research.
