The ARHGAP35 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout population derived from the HT29 human colorectal adenocarcinoma epithelial cell line. This model provides targeted disruption of the ARHGAP35 gene, encoding p190-A RhoGAP, a key regulator of Rho GTPase signaling, actin dynamics, cell migration, and adhesion. The polyclonal format ensures stable gene inactivation while maintaining population heterogeneity, making it a versatile tool for studying tumor-relevant signal transduction pathways.
HT29 is a widely used colorectal adenocarcinoma line harboring mutations in APC, TP53, and KRAS, recapitulating major oncogenic drivers of sporadic colorectal cancer. Established from a primary tumor of a 44-year-old female, these epithelial cells retain the ability to produce mucin and serve as a critical model for intestinal tumor biology, including studies on tumorigenesis, metastatic progression, and therapeutic responses. Their genetic background sensitizes them to altered Rho GTPase activity, enabling dissection of cooperative signaling networks.
ARHGAP35 encodes p190-A RhoGAP, which inactivates RhoA, Rac1, and Cdc42 by stimulating GTP hydrolysis, thereby reducing actin stress fiber formation, focal adhesion turnover, and cellular contractility. Its activity is regulated by upstream factors such as EGF, PDGF, Src kinases, and integrin engagement via FAK. At focal adhesions, p190-A interacts with p120 catenin, Cortactin, and tensin, and its phosphorylation by Src couples extracellular cues to cytoskeletal reorganization. Downstream targets include the GTP-bound forms of RhoA, Rac1, and Cdc42, linking p190-A to suppression of cell migration and invasion.
In the HT29 context, ARHGAP35 knockout leads to hyperactivation of Rho GTPases, resulting in enhanced stress fiber assembly, strengthened focal adhesions, and increased migratory and invasive capacity. Given the coexisting APC, TP53, and KRAS mutations, loss of p190-A may cooperate with these oncogenic pathways to promote aggressive tumor cell behavior. This derivative cell model is therefore invaluable for investigating how Rho GTPase hyperactivity intersects with known colorectal cancer drivers to influence epithelial-mesenchymal transition, 3D invasion, and drug sensitivity.
These knockout cells are ideally suited for a range of advanced research applications, including RhoA/Rac1 activation-GST pull-down assays, western blotting for ARHGAP35 and downstream effectors, phalloidin staining of actin stress fibers, and transwell migration/invasion experiments. They enable high-content screening of inhibitors blocking Rho GTPase or Src/FAK signaling, as well as mechanistic studies of colorectal cancer metastasis and cytoskeletal dynamics. The cells offer a reliable platform for both basic and translational investigations. For further information, please contact Ascent Research.
