The ILKAP Knockout HT29 Polyclonal Cells consist of a CRISPR/Cas9-edited polyclonal knockout cell population in the human HT29 colorectal adenocarcinoma cell line. This heterogeneous pool of gene-disrupted cells avoids clonal bias and provides a robust loss-of-function model for studying ILKAP function. The CRISPR/Cas9-mediated gene disruption effectively abolishes ILKAP protein expression, enabling reproducible functional studies.
HT29 cells, derived from a primary colorectal adenocarcinoma of a 44-year-old female, are an established epithelial model for colorectal cancer research. They retain active Wnt/??-catenin and PI3K/AKT pathways and are widely used for investigating epithelial differentiation, barrier function, and oncogenic signaling. Their well-characterized behavior makes them ideal for knockout-based analyses.
ILKAP encodes a serine/threonine phosphatase that directly dephosphorylates integrin-linked kinase (ILK), thereby attenuating ILK-mediated signaling. Under physiological conditions, ILKAP is transcriptionally regulated by p53 and activated by integrin engagement and growth factors such as EGF and TGF-??. Within focal adhesion complexes, ILKAP interacts with ILK, PINCH, and parvin, and its catalytic activity suppresses ILK-dependent phosphorylation of AKT and GSK-3??. Consequently, loss of ILKAP results in hyperactive ILK signaling, leading to elevated phospho-AKT and phospho-GSK-3?? levels, which promote cell proliferation, survival, and actin cytoskeletal remodeling. This signaling axis integrates upstream inputs from FAK and Src and influences ??-catenin stability, coupling adhesion-dependent signals to transcriptional outputs.
In HT29 colorectal cancer cells, ILKAP is predicted to function as a tumor suppressor. Its knockout leads to unchecked ILK-AKT-GSK-3?? signaling that can further enhance ??-catenin activity, synergizing with the intrinsic Wnt pathway dysregulation in these cells. This polyclonal knockout model thus provides a relevant platform to dissect how phosphatase regulation of integrin signaling impacts colon cancer cell adhesion, migration, survival, and response to therapeutic agents.
Typical applications include western blot analysis of phospho-AKT (Ser473) and phospho-GSK-3?? (Ser9) to validate pathway activation, RT-qPCR for downstream gene targets, and functional assays such as MTT proliferation, Annexin V apoptosis, and transwell migration/invasion. Immunofluorescence for focal adhesion markers and flow cytometric cell cycle analysis further enable detailed phenotypic characterization. This product supports elucidation of ILKAP??s tumor suppressor role, drug sensitivity screens, and exploration of integrin-ILK signaling networks. For further information, please contact Ascent Research.
