GXYLT2 Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from HT29 human colorectal adenocarcinoma epithelial cells, engineered for targeted disruption of the GXYLT2 gene. This loss-of-function model enables investigation of xylosyltransferase activity critical for matriglycan chain biosynthesis on alpha-dystroglycan. The polyclonal nature of the knockout pool reflects a heterogeneous collection of edited alleles generated by non-homologous end joining following Cas9-mediated double-strand breaks, providing a comprehensive gene perturbation tool without single-cell cloning.
The HT29 parental cell line is an extensively characterized model of human colorectal adenocarcinoma, originally isolated from a 44-year-old female patient. These epithelial cells retain differentiated features of intestinal epithelium and are widely used to study colorectal cancer biology, including proliferation, differentiation, and metastatic behavior. Additionally, HT29 cells serve as a platform for examining intestinal epithelial transport and barrier function, making them particularly relevant for evaluating the role of glycosylation pathways in disease contexts.
GXYLT2 encodes a glycosyltransferase that adds xylose to glucosylated O-mannose on alpha-dystroglycan (DAG1), a requisite step for matriglycan polymer extension. This modification is essential for dystrophin-glycoprotein complex assembly and laminin binding. The enzyme functions downstream of initial O-mannosylation by POMT1/POMT2 and subsequent POMGnT1/B3GALNT2 actions, cooperating with GXYLT1 to prime the glycan for LARGE1-mediated elongation. TMEM5 and Fukutin also contribute to this biosynthetic network. Loss of GXYLT2 disrupts dystroglycan glycosylation, impairing cell adhesion and complex stability, with downstream effects on cytoskeletal linkage and signal transduction.
In HT29 colorectal adenocarcinoma cells, GXYLT2 knockout provides a physiologically relevant system to dissect the intersection of glycosylation and cancer cell behavior. Dysregulation of dystroglycan glycosylation has been implicated in tumor progression, altering cell-matrix interactions that govern migration and invasion. This model enables examination of how matriglycan deficiency modulates epithelial adhesion dynamics, anoikis resistance, and metastatic potential. Moreover, because dystroglycanopathy-associated mutations lead to muscular dystrophy and brain malformations, the model can be applied to study disease mechanisms in an epithelial context, complementing muscle- or neuron-specific systems.
Typical applications include probing GXYLT2-dependent glycosylation via IIH6 immunofluorescence or laminin overlay assays to detect functional dystroglycan. Western blotting and RT-qPCR confirm knockout and assess transcriptional effects, while cell adhesion, migration, and invasion assays quantify phenotypic changes. Flow cytometry for surface dystroglycan complements biochemical analyses. This product also supports screening for glycosylation modulators or evaluation of gene therapy strategies for dystroglycanopathies. For further information, please contact Ascent Research.
