The AAGAB Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population in the HT29 human colorectal adenocarcinoma line, targeting the AAGAB gene. This heterogeneous loss-of-function model enables investigation of the alpha- and gamma-adaptin binding protein (AAGAB) in clathrin-mediated endocytosis and receptor internalization within a colorectal cancer epithelial context.
HT29 cells, derived from a human colorectal adenocarcinoma, are a tumorigenic epithelial line widely used for intestinal biology, colorectal cancer, and drug absorption studies. Their polarized epithelial features suit membrane trafficking and receptor-mediated process investigations, making them an appropriate host for assessing endocytic dysfunction due to AAGAB knockout.
At the molecular level, AAGAB acts as an adaptor that binds alpha-adaptin subunits of AP-2 (AP2A1, AP2A2) and gamma-adaptin of AP-1 (AP1G1), as well as clathrin heavy chain, to promote clathrin coat assembly. This facilitates internalization of cargoes like EGFR and transferrin receptor. Upstream, AP-2/AP-1 and cargo receptors initiate coat nucleation. Disrupted AAGAB expression impairs coated pit formation, reducing uptake of these receptors and attenuating downstream signaling cascades.
In the HT29 colorectal adenocarcinoma context, loss of AAGAB-mediated coat assembly provides a model to dissect endocytic dysfunction in cancer. The tumorigenic background enables study of sustained receptor tyrosine kinase activity and altered nutrient transporter traffic. Additionally, the model relates to punctate palmoplantar keratoderma, associated with AAGAB mutations, offering a platform to examine endocytic defects relevant to this rare disorder.
These polyclonal knockout cells support diverse applications, including transferrin uptake and EGFR internalization assays to quantify endocytosis, immunofluorescence for clathrin and AP-2 to visualize coat defects, Western blotting for receptor levels, and cell proliferation assays to assess functional impact. Transcriptomic profiling via RNA-seq can uncover compensatory pathways. This model facilitates mechanistic studies of trafficking in colorectal cancer and related diseases. For details, contact Ascent Research.
