The ACAP2 Knouckout HT29 Polyclonal Cells consist of a CRISPR/Cas9-edited polyclonal knockout population with disrupted ACAP2 in the HT29 colorectal adenocarcinoma line. This heterogeneous pool enables robust loss-of-function studies while reducing clonal selection bias, making it suitable for investigating complex phenotypes like migration and invasion in a tumor-relevant context.
HT29 cells, derived from a primary colorectal adenocarcinoma from a 44-year-old Caucasian female, serve as a well-characterized intestinal epithelial model with an absorptive enterocyte-like phenotype. Widely used in colorectal cancer research, these cells form polarized monolayers and facilitate studies on barrier function, signal transduction, and tumor cell behavior. In the ACAP2 knockout setting, HT29 provides a clinically pertinent background to examine ARF6-driven trafficking in colorectal cancer progression.
ACAP2 encodes a GTPase-activating protein for ARF6, catalyzing GTP hydrolysis to inactivate ARF6 and negatively regulate integrin recycling. Its activity is governed by membrane phosphoinositides like PI(4,5)P2 and PI(3,4,5)P3, and inputs from receptor tyrosine kinases such as EGFR. ACAP2 interacts with ARF6 and BAR domain proteins, and its GAP function suppresses downstream effectors including Rac1 and phospholipase D. Loss of ACAP2 leads to sustained ARF6 activation, enhanced recycling of adhesion receptors, and increased cell migration and invasion.
In HT29 colorectal adenocarcinoma cells, ACAP2 knockout yields unchecked ARF6 activation, augmenting integrin recycling and altering adhesion dynamics to promote motility and invasion??key processes in metastasis. This model allows dissection of the ACAP2-ARF6 regulatory axis within an intestinal epithelial context, offering insights into how disrupted endosomal trafficking fuels tumor progression while maintaining some differentiated epithelial features.
Applications include studying ARF6 signaling in cancer metastasis, integrin trafficking, and pathway-targeted drug screening. Representative assays encompass Western blot for knockout validation and ARF6-GTP levels, Transwell migration/wound healing, integrin recycling via antibody uptake, immunofluorescence, adhesion assays, and phospho-signaling analysis of Akt and Erk. RT-qPCR can monitor ARF6 target genes. For further technical details, please contact Ascent Research.
