The ATP11A Knockout HT29 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population derived from human HT29 colorectal adenocarcinoma epithelial cells, providing a loss-of-function model for the ATP11A gene. The polyclonal composition ensures heterogeneous editing events across the population, enabling robust, population-level phenotypic analyses without clonal selection bias. CRISPR/Cas9-mediated disruption abrogates functional ATP11A protein expression, offering a versatile tool for dissecting phospholipid asymmetry, membrane dynamics, and associated cellular processes in a cancer-relevant background.
HT29 is a widely studied human colorectal adenocarcinoma cell line with epithelial morphology, used in cancer research, drug screening, and intestinal biology. Its adherent growth and signaling networks make it ideal for studying colorectal cancer progression and membrane trafficking.
ATP11A encodes a P4-ATPase flippase that, in obligate complexation with the chaperone TMEM30A (CDC50A), actively translocates phosphatidylserine and phosphatidylethanolamine from the exoplasmic to the cytoplasmic membrane leaflet. This vectorial lipid transport sustains plasma membrane phospholipid asymmetry. The flippase is regulated by TMEM30A, cellular stress, and membrane lipid composition. Functionally, ATP11A suppresses aberrant phosphatidylserine externalization to prevent unwanted apoptotic cell clearance signals, modulates Rho GTPase signaling to control actin-dependent cell migration and adhesion, and facilitates endocytic cargo trafficking through membrane curvature regulation. Interaction with TMEM30B further extends flippase functionality to organelle membranes. Disruption of ATP11A thus collapses phospholipid asymmetry, leading to sustained phosphatidylserine exposure and altered vesicle trafficking.
In the colorectal cancer context of HT29 cells, ATP11A knockout disrupts membrane asymmetry, resulting in increased cell surface phosphatidylserine. This may mimic early apoptotic signals or modulate immune cell interactions, thereby influencing tumor immune evasion and metastatic potential. As colorectal cancer progression requires dynamic remodeling of lipid distribution for migration and invasion, this model enables dissection of how flippase activity governs motility, endocytosis, and Rho GTPase-dependent signaling. Moreover, the exposure of phosphatidylserine serves as a ligand for phagocytic receptors and affects microvesicle shedding, allowing exploration of how colorectal tumor cells manipulate phospholipid scrambling to sculpt the tumor microenvironment.
Research applications encompass investigation of phospholipid asymmetry in colorectal cancer, functional analysis of flippase-dependent cell migration and invasion, and screening for pharmacological modulators of membrane asymmetry pathways. Representative assays include annexin V staining by flow cytometry or immunofluorescence to quantify phosphatidylserine exposure, transwell migration and invasion assays, transferrin uptake analysis for endocytosis, Western blotting for ATP11A expression, confocal imaging of flippase complex localization, and RT-qPCR for transcript validation. These polyclonal knockout cells also provide a platform for immune evasion studies, given the impact of aberrant phosphatidylserine on immune recognition. For further information and ordering, please contact Ascent Research.
