The ARK2N Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 colorectal adenocarcinoma cell line, with targeted disruption of ARK2N. This heterogeneous pool captures a range of knockout efficiencies, enabling bulk functional analyses without clonal selection bias. The model is suited for investigating ARK2N roles in lysosomal biology, mTORC1 signaling, and autophagy within an intestinal epithelial cancer context.
The parental HT29 line was established from a primary colon adenocarcinoma of a 44-year-old female. HT29 cells carry mutations in APC and TP53, recapitulating key genetic lesions of colorectal cancer. They serve as a well-characterized model of intestinal epithelial physiology and are widely used for studying oncogenic signaling, barrier function, and drug responses. Their adherent morphology and robust growth facilitate generation of gene-edited derivatives to dissect colon carcinogenesis.
ARK2N encodes a lysosomal adaptor protein that physically bridges the BORC complex to kinesin motors KIF5B and KIF3A, facilitating anterograde lysosomal transport to the cell periphery. Peripheral lysosome positioning suppresses mTORC1 activity and promotes autophagy. ARK2N interacts with BORC subunits (BLOC1S1, BLOC1S2) and ARL8B to regulate motor coupling. Downstream, it modulates mTORC1 phosphorylation of S6K and 4E-BP1, and influences autophagic markers LC3-II and p62. The pathway responds to nutrient availability and growth factors.
In HT29 colorectal cancer cells, ARK2N knockout induces perinuclear lysosome clustering, mTORC1 hyperactivation, and reduced autophagic flux. Since mTORC1 hyperactivity and autophagy suppression promote tumorigenesis, this model helps dissect lysosomal trafficking contributions to cancer cell growth and metabolism. The APC and TP53 mutant background further allows exploration of crosstalk between lysosomal positioning and Wnt/p53 pathways, suggesting vulnerabilities for therapeutic targeting.
Applications include studying lysosomal dynamics in colorectal cancer, mTORC1 signaling, autophagy regulation, and drug sensitivity. Assays with this polyclonal knockout population encompass western blotting for p-S6K and p-4E-BP1, LC3 and p62 turnover, LAMP1 immunofluorescence, Lysotracker red staining, kinesin co-immunoprecipitation, and cell migration/proliferation assays. Thus, these cells support robust phenotypic interrogation in a genetically defined adenocarcinoma model. For further details, please contact Ascent Research.
