ASPH Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population derived from the HT29 colorectal adenocarcinoma cell line, featuring targeted disruption of the aspartate beta-hydroxylase (ASPH) gene. This gene-edited tool enables loss-of-function studies of ASPH in a colorectal cancer context, supporting investigations into Notch and calcium signaling networks central to tumor biology.
The parental HT29 cell line originates from a primary colorectal adenocarcinoma of a female patient and serves as an established model for colorectal cancer research. These epithelial cells maintain oncogenic pathways characteristic of colorectal tumors, including dysregulated Wnt and MAPK signaling, and are therefore appropriate for interrogating ASPH function in a disease-relevant setting.
ASPH hydroxylates aspartate and asparagine residues in EGF-like domains of Notch receptors and ligands, a modification critical for efficient Notch activation. Upon ligand binding, the Notch intracellular domain (NICD) is released, translocates to the nucleus, and forms a complex with RBP-J to induce transcription of target genes such as HES1. ASPH also binds calmodulin and facilitates calcium entry via TRPC3 channels, linking growth factor signals (EGF, TGF-??) to calcium homeostasis and Notch signaling. Disruption of ASPH is thus expected to attenuate NICD generation, HES1 expression, and calcium influx, impairing these interconnected pathways.
In HT29 cells, ASPH contributes to malignant traits like proliferation and epithelial-mesenchymal transition. The polyclonal knockout population circumvents clonal artifacts, allowing assessment of averaged phenotypic effects. Researchers can use this model to dissect how loss of ASPH-dependent Notch potentiation and calcium modulation limits HT29 tumorigenic capacity, revealing crosstalk between Notch and Wnt/EGFR pathways that sustain colorectal cancer progression.
These cells are compatible with a variety of biochemical and functional assays. Western blotting and RT-qPCR enable quantification of pathway markers (NICD, HES1), while calcium imaging measures altered calcium dynamics. MTT, colony formation, and wound healing assays evaluate viability, clonogenicity, and migration. Notch reporter assays directly gauge pathway activity. Additionally, this model suits drug screening for inhibitors targeting ASPH-related signaling. For technical inquiries, contact Ascent Research.
