ALKBH4 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population generated in the HT29 human colorectal adenocarcinoma background. This product provides a loss-of-function model for ALKBH4, a lysine demethylase that regulates actin polymerization and cell motility. The polyclonal nature ensures a heterogeneous pool with disrupted gene function, avoiding clonal artifacts and enabling robust phenotypic screening. CRISPR/Cas9-mediated gene disruption eliminates functional ALKBH4, facilitating investigation of its role in cytoskeletal dynamics and metastatic behavior.
The HT29 cell line, originally derived from a 44-year-old Caucasian female with colorectal cancer, is a well-characterized epithelial model extensively used in intestinal physiology, cancer biology, and drug absorption studies. These cells retain epithelial morphology and key signaling pathways, and exhibit a capacity to undergo epithelial-mesenchymal transition (EMT) upon appropriate stimulation. As a mesenchymally competent adenocarcinoma line, HT29 provides an optimal host for studying genes implicated in tumor invasion and metastasis.
ALKBH4 functions as an actin demethylase, specifically targeting monomethylated lysine-84 (K84me1). This demethylation event promotes actin filament polymerization, ARP2/3 complex-mediated branching, and cofilin-dependent severing, thereby enhancing actomyosin contractility through phosphorylation of myosin light chain. These processes drive cell migration and invadopodia formation. ALKBH4 expression is regulated by EMT-associated transcription factors such as SNAI1 and TWIST1 and operates downstream of RhoA/ROCK signaling. It interacts with actin, myosin, ARP2/3, and cofilin, orchestrating cytoskeletal remodeling for invasive motility.
In the HT29 colorectal adenocarcinoma context, knockout of ALKBH4 disrupts actin dynamics, leading to impaired cell migration, invasion, and invadopodia-mediated extracellular matrix degradation. Since HT29 cells are capable of undergoing EMT, this model captures a critical step in colorectal cancer progression. The loss of ALKBH4-mediated actin demethylation attenuates actomyosin contractility and reduces metastatic potential, providing a physiologically relevant platform to dissect the molecular interplay between post-translational modifications of actin and aggressive tumor cell behavior.
These polyclonal knockout cells are ideally suited for transwell migration and Matrigel invasion assays, immunofluorescence of actin cytoskeleton and invadopodia markers, co-immunoprecipitation of ALKBH4 with actin/myosin complexes, western blotting for actin K84me1 and phospho-myosin light chain, and RT-qPCR analysis. Applications encompass dissecting actin methylation-dependent signaling in colorectal cancer metastasis and screening small-molecule inhibitors. For technical inquiries or bulk orders, please contact Ascent Research.
