The ATG3 Knockout DLD-1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the human DLD-1 colorectal adenocarcinoma cell line, engineered for loss-of-function studies of the autophagy-related gene ATG3. This knockout model disrupts the ATG3 locus via CRISPR/Cas9-mediated gene disruption, generating a heterogeneous pool of edited cells suitable for functional analyses without single-cell clonal selection. The polyclonal format preserves population-level heterogeneity while eliminating target gene function, providing a robust tool for investigating autophagy-dependent processes.
The DLD-1 cell line originates from a Duke’s type C colorectal adenocarcinoma in a 45-year-old male and carries well-characterized mutations in APC, TP53, KRAS, and PIK3CA, making it a representative model for colorectal cancer studies. These epithelial cells exhibit features relevant to tumor progression, including dysregulated proliferation, invasion, and signaling pathways. The genetic background of DLD-1 provides a context for examining how autophagy modulation impacts oncogenic signaling and therapeutic responses.
ATG3 encodes an E2-like enzyme that catalyzes the covalent conjugation of ATG8 family proteins (MAP1LC3A, MAP1LC3B, MAP1LC3C, GABARAP, GABARAPL1, GABARAPL2) to phosphatidylethanolamine on autophagosome membranes, a critical step in autophagosome biogenesis and maturation. This activity requires prior activation by the E1-like enzyme ATG7 and the ATG12-ATG5-ATG16L1 conjugation system, which acts as an E3-like ligase. ATG3 function is regulated upstream by the ULK1 complex, mTOR and AMPK signaling, and stress stimuli such as starvation and hypoxia. Through its role in LC3 lipidation, ATG3 directly controls autophagic flux, impacting downstream processes such as cargo degradation and recycling. It interacts closely with ATG7, the ATG12-ATG5 conjugate, ATG16L1, and ATG8 family members.
In the context of DLD-1 colorectal cancer cells, ATG3 knockout impairs autophagic flux, compromising the cellular response to nutrient deprivation and chemotherapeutic stress. Given that autophagy can have pro-survival or pro-death functions depending on the tumor microenvironment, disrupting ATG3 provides a means to dissect these dual roles in colorectal cancer. This model is particularly relevant for studying chemoresistance mechanisms, as autophagy often protects cancer cells against drug-induced apoptosis. The DLD-1 ATG3 knockout cells enable the interrogation of autophagy-dependent signaling networks and the identification of vulnerabilities that can be targeted therapeutically.
Researchers can utilize the ATG3 Knockout DLD-1 Polyclonal Cells for a variety of experimental approaches, including Western blot analysis of ATG3, LC3-II, and p62/SQSTM1 to assess autophagic flux, immunofluorescence detection of LC3 puncta, and cell viability assays under starvation or chemotherapeutic treatments. These cells are also suitable for screening autophagy-modulating compounds and exploring non-autophagic functions of ATG3. We recommend verifying target gene disruption via genomic PCR and Sanger sequencing, and confirming functional loss through autophagic flux assays in the presence of lysosomal inhibitors such as chloroquine. For technical inquiries or ordering details, please contact Ascent Research.
