The GNPDA2 Knockout HT29 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout population of the human colorectal adenocarcinoma HT29 cell line, engineered to disrupt the GNPDA2 gene. This polyclonal cell product provides a heterogeneous pool of edited alleles, enabling robust loss-of-function studies without clonal selection artifacts. The knockout model is generated via CRISPR/Cas9-mediated gene disruption, resulting in abrogation of functional GNPDA2 protein expression. This tool is designed for researchers investigating hexosamine metabolism and its intersection with oncogenic signaling and metabolic disorders.
The HT29 host cell line, derived from a primary human colorectal adenocarcinoma, serves as a well-characterized model of intestinal epithelial biology. HT29 cells harbor mutations in the tumor suppressors p53 and APC, facilitating studies of colorectal cancer initiation and progression. These cells are extensively used to examine intestinal absorption, barrier function, and epithelial polarity. The combination of these genetic lesions with a targeted disruption of GNPDA2 allows interrogation of metabolite-driven signaling in a relevant oncogenic background.
GNPDA2 (Glucosamine-6-Phosphate Deaminase 2) catalyzes the deamination of glucosamine-6-phosphate to fructose-6-phosphate, connecting amino sugar catabolism to glycolysis. It integrates into the hexosamine pathway, influencing UDP-GlcNAc synthesis and protein O-GlcNAcylation. Upstream regulators include mTORC1 signaling, c-Myc, and HIF1??, while downstream outputs involve glycolytic flux and O-GlcNAc modifications. The enzyme forms homodimers and acts on glucosamine-6-phosphate, with pathway partners GFAT, GNPNAT1, and hexokinase.
In HT29 cells, which exhibit dysregulated Wnt and p53 pathways due to APC and p53 mutations, GNPDA2 perturbation is particularly consequential. Given its association with obesity, colorectal cancer, and metabolic syndrome, this knockout model enables dissection of how altered hexosamine flux modulates oncogenic signaling and metabolic reprogramming. The polyclonal nature of the edited pool provides a more physiological representation of heterogeneous tumor cell populations, making it suitable for studying clonal variation in metabolic adaptation and therapeutic response.
This product is ideally suited for metabolic flux analyses using Seahorse assays and LC-MS-based metabolite profiling, as well as for investigating O-GlcNAcylation dynamics via western blotting and immunoprecipitation. Researchers can employ these cells in proliferation, apoptosis, and migration assays to assess functional consequences of GNPDA2 loss. The model further supports RT-qPCR and RNA sequencing for downstream target validation, including glycolytic enzymes and O-GlcNAc-modified proteins. For additional technical details or to place an order, please contact Ascent Research.
