The GNPDA1 Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal cell population derived from the HT29 colorectal adenocarcinoma line, designed for constitutive disruption of GNPDA1. This knockout model enables investigation of GNPDA1 loss on hexosamine pathway flux and glycosylation. The polyclonal format preserves population diversity from heterogeneous edits, allowing study of overall gene function without clonal bias, suited for cancer metabolism and signal transduction research.
The HT29 host cell line was established from a primary colorectal adenocarcinoma of a 44-year-old Caucasian female. Serving as a well-characterized model of intestinal epithelial cells, HT29 cells are widely employed in colorectal cancer research to dissect oncogenic signaling, metabolic reprogramming, and therapeutic responses. Their epithelial morphology and ability to form polarized monolayers make them relevant for studying intestinal biology and tumor-associated glycosylation changes.
GNPDA1 encodes glucosamine-6-phosphate deaminase 1, converting glucosamine-6-phosphate to fructose-6-phosphate and ammonia, diverting substrate from UDP-GlcNAc synthesis. In the hexosamine biosynthetic pathway, GNPDA1 is a critical node regulated by MYC, HIF1A, and OGT, interacting with GFPT1 and GNPNAT1. GNPDA1 loss increases hexosamine flux toward UDP-GlcNAc, elevating O-GlcNAcylation by OGT and altering glycosyltransferase-mediated glycosylation. This couples nutrient sensing to post-translational modification, impacting signal transduction and metabolic control.
In HT29 colorectal cancer cells, GNPDA1 knockout disrupts hexosamine intermediate balance, altering O-GlcNAcylation and metabolic rewiring affecting proliferation, survival, and invasion. This model investigates how GNPDA1-dependent glycosylation intersects with colorectal tumorigenesis, potentially revealing therapeutic vulnerabilities. It also provides a platform for studying congenital disorders of glycosylation.
Researchers can use these knockout cells for metabolic flux analysis with 13C-glucose, UDP-GlcNAc LC-MS, and O-GlcNAcylation Western blot or lectin blot. Functional assays like MTS proliferation, colony formation, annexin V apoptosis, and migration assays characterize GNPDA1 loss phenotypes. RNA-seq can profile transcriptomic changes. Applications include colorectal cancer metabolism, hexosamine pathway targeting, O-GlcNAcylation dynamics, and drug target validation. For technical information, contact Ascent Research.
