The ATG7 Knockout HT29 Polyclonal Cells consist of a CRISPR/Cas9-edited polyclonal population of HT29 colorectal adenocarcinoma cells with targeted ATG7 disruption. This heterogeneous pool carries diverse loss-of-function mutations, providing a robust model for studying autophagy-dependent processes in intestinal epithelial cancer cells. The polyclonal format retains natural genetic variability while ensuring reliable loss of ATG7 function across the cell population.
HT29 cells were isolated from a primary colon adenocarcinoma in a 44-year-old female and are widely used as an intestinal epithelial model for colorectal cancer research. These adherent cells exhibit epithelial morphology and harbor mutations in APC and TP53, enabling studies of colon carcinoma biology and drug responses in a relevant genetic context.
The ATG7 gene encodes an E1-like ubiquitin-activating enzyme that plays an essential role in autophagy by catalyzing two ubiquitin-like conjugation reactions. ATG7 activates ATG12 and facilitates its transfer to ATG10, ultimately leading to covalent linkage between ATG12 and ATG5; this ATG12?CATG5 conjugate then forms a complex with ATG16L1 and functions as an E3-like ligase for LC3 lipidation. In parallel, ATG7 also activates LC3 and transfers it to ATG3, enabling its conjugation to phosphatidylethanolamine to generate lipidated LC3-II, a key component of autophagosomal membranes. ATG7 is regulated by upstream nutrient and energy sensors, including mTORC1 inhibition and AMPK activation, which signal through the ULK1 complex and PI3KC3?CBeclin-1 axis to initiate autophagy. Downstream, loss of ATG7 prevents LC3 lipidation and ATG5?CATG12 conjugate formation, blocking autophagosome biogenesis and impairing autophagic flux, which can be monitored by accumulation of p62 and diminished LC3-II turnover.
In HT29 colorectal adenocarcinoma cells, ATG7 knockout dissects autophagy’s role in tumor survival, drug resistance, and metastasis. Colorectal tumors often rely on autophagy for stress adaptation; ATG7 loss can sensitize cells to apoptosis and reveal therapeutic vulnerabilities. This model also enables study of autophagy crosstalk with Wnt/??-catenin and KRAS pathways, and serves for host-pathogen interaction studies and autophagy modulator screening in a disease-relevant cell system.
Typical assays include western blot for LC3-II and p62, fluorescence microscopy for LC3 puncta, autophagic flux assays with chloroquine, cell viability and apoptosis flow cytometry, and co-immunoprecipitation for ATG7?CATG12 interaction. These cells support autophagy-related drug screening and CRISPR-based double-knockout studies. For further information or custom cell engineering inquiries, please contact Ascent Research.
