The GPT2 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colorectal adenocarcinoma cell line. This product features targeted disruption of the GPT2 gene via CRISPR/Cas9, generating a heterogeneous loss-of-function pool ideal for studying glutamic pyruvate transaminase 2 function without clonal selection. The polyclonal format ensures representation of diverse genetic alterations, avoiding the limitations of single-cell clones.
HT29 is a widely used human epithelial colorectal adenocarcinoma cell line originally established from a primary tumor of a 44-year-old Caucasian female. These cells serve as a robust model of intestinal epithelium, exhibiting stable epithelial morphology and retaining key characteristics of colorectal cancer, including metabolic reprogramming. HT29 cells are extensively applied in studies of intestinal biology, drug transport, and oncogenic signaling, providing a highly relevant cellular context for gene-editing approaches.
GPT2 encodes the mitochondrial enzyme glutamic pyruvate transaminase 2, which catalyzes the reversible transamination of alanine and 2-oxoglutarate to pyruvate and glutamate. This reaction bridges amino acid metabolism with gluconeogenesis and the TCA cycle. GPT2 expression is regulated by PPARGC1A and FOXO1 downstream of glucagon and cAMP signaling, and it responds to glucocorticoids. Functionally, GPT2 interacts with GLUD1, GOT2, MDH2, and the PDH complex, forming a metabolic network that controls pyruvate and glutamate flux. Thus, GPT2 is critical for alanine-driven anaplerosis, nitrogen metabolism, and maintenance of amino acid and redox homeostasis.
In HT29 colorectal cancer cells, GPT2 knockout abolishes alanine transamination, depleting pyruvate and glutamate pools and severely impairing TCA cycle anaplerosis, glutathione synthesis, and nucleotide biosynthesis. This metabolic disruption sensitizes cells to nutrient stress and highlights GPT2-dependent vulnerabilities in colorectal cancer, where high anaplerotic flux supports proliferation. The model also reflects GPT2 deficiency-linked neurodevelopmental disorders, underscoring the enzyme’s role across tissues.
These GPT2 knockout polyclonal cells enable detailed metabolic flux analyses using [13C]-alanine tracing, Seahorse respirometry, and metabolomics profiling to quantify perturbations in central carbon metabolism. They support investigation of anaplerotic pathways, gluconeogenic activity, and mitochondrial function through alanine transaminase assays, mitochondrial mass measurement, and glutathione quantification. The cells are suited for cancer metabolism research, drug target discovery, and resistance screening in colorectal cancer models. For further information, please contact Ascent Research.
