The ATP11B Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colorectal adenocarcinoma epithelial cell line, featuring targeted disruption of the ATP11B gene. This loss-of-function model enables researchers to investigate the consequences of ATP11B ablation in a colon cancer context without clonal selection biases.
HT29 cells are a well-established human colorectal adenocarcinoma cell line with epithelial morphology, originally isolated from a primary tumor of a 44-year-old female. These cells are extensively used as a model system for studying intestinal epithelial barrier function, drug absorption, and the molecular mechanisms underlying colorectal adenocarcinoma progression.
ATP11B encodes a P4-ATPase flippase that translocates phosphatidylserine (PS) and phosphatidylethanolamine (PE) to the cytoplasmic leaflet, preserving membrane asymmetry. Its activity is regulated by MYC, TP53, and the CDC50 family (TMEM30A/B), and is modulated by calcium and protein kinase C. ATP11B interacts with CDC50A/B to form a functional complex. Knockout of ATP11B results in abnormal PS externalization, an ‘eat-me’ signal for TAM receptors (Tyro3, Axl, MerTK) that triggers phagocytic clearance. Downstream, the AKT pathway, Rac1 GTPase, and focal adhesion turnover are affected, disrupting cell migration and apoptotic signaling.
In HT29 colorectal adenocarcinoma cells, disruption of ATP11B alters phospholipid asymmetry, affecting membrane dynamics, endocytic recycling, and exosome biogenesis. This model is pivotal for studying how aberrant PS exposure influences immune evasion and tumor microenvironment interactions. The polyclonal knockout population retains genetic heterogeneity, enabling robust assessment of population-level phenotypes in barrier function or drug response assays.
Key applications include flow cytometric detection of surface PS with Annexin V, Transwell migration assays to evaluate motility changes, and macrophage phagocytosis assays to study efferocytosis. Apoptosis assays, Western blotting, RT-qPCR, and immunofluorescence can confirm knockout and analyze signaling. TEER measurement assesses barrier integrity, and lipidomics reveals phospholipid alterations. These tools support research into colorectal cancer lipid metabolism, PS-mediated immune evasion, metastasis, and flippase inhibitor screening. For more information, contact Ascent Research.
