The ATG4C Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HAP1 human chronic myeloid leukemia (CML) cell line, designed to disrupt the ATG4C gene. This loss-of-function model provides a heterogeneous pool of cells for studying ATG4C-dependent autophagy processes in a cancer-relevant background.
HAP1 is a near-haploid human male CML-derived cell line, BCR-ABL positive, and widely used for gene knockout studies due to its tractable genome. As a leukemia model, HAP1 cells retain hematopoietic malignancy features, enabling investigation of autophagy in tumor cell survival and drug resistance.
ATG4C encodes a cysteine protease that cleaves the C-termini of pro-LC3 and GABARAP family proteins, exposing glycine for phosphatidylethanolamine (PE) conjugation, a key step in LC3-II formation and autophagosome maturation. ATG4C is regulated upstream by mTORC1 and AMPK in response to nutrient deprivation and ULK1 kinase activity, while p53 modulates its expression under stress. Downstream, ATG4C processes LC3A, LC3B, GABARAP, GABARAPL1, and GABARAPL2, interacting with ATG7 and ATG3 in the ubiquitin-like conjugation system. Additional interplay with BCL2 and Beclin-1 connects autophagy to apoptosis.
In HAP1 cells, ATG4C knockout disrupts LC3 lipidation and impairs autophagic flux, leading to accumulation of substrates like p62/SQSTM1. This defect compromises the cytoprotective autophagy that supports leukemia cell survival under metabolic stress or chemotherapy, making the model valuable for exploring autophagy-targeting therapeutic strategies.
Applications include Western blotting for LC3 lipidation, autophagy flux assays with bafilomycin A1, and fluorescence-based reporters (GFP-LC3-RFP) to monitor autophagosome formation. The cells are suited for drug sensitivity profiling, apoptosis assays (Annexin V) to examine autophagy-apoptosis crosstalk, and screening for ATG4C-selective inhibitors. Researchers can also investigate neurodegenerative or infectious disease mechanisms involving autophagy. For additional information, please contact Ascent Research.
