The ABAT Knockout HCT 116 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population with targeted disruption of the human ABAT gene in the HCT 116 colorectal carcinoma cell line. This loss-of-function model abolishes 4-aminobutyrate aminotransferase (GABA transaminase) enzymatic activity, providing a genetically defined system to investigate GABA metabolism and its intersection with central carbon metabolism in cancer. The polyclonal nature reflects a heterogeneous collection of edited cells generated by Cas9-mediated gene disruption without single-cell cloning, enabling pooled functional studies.
The parental HCT 116 cell line is an established model of colorectal carcinoma derived from a male adult patient, exhibiting epithelial morphology, microsatellite instability-high (MSI-H) status, and a KRAS G13D activating mutation. These cells are widely employed in cancer biology for studying tumorigenesis, drug responses, and metabolic reprogramming. Their genetic background, including the KRAS mutation, makes them particularly relevant for investigating oncogene-driven metabolic adaptations and the role of the GABA shunt in sustaining proliferation under nutrient stress.
ABAT encodes the mitochondrial enzyme GABA transaminase, which depends on pyridoxal phosphate (PLP) as a cofactor and catalyzes the conversion of GABA and ??-ketoglutarate to succinic semialdehyde and glutamate. Functionally, ABAT sits at a metabolic hub linking the GABA shunt to the tricarboxylic acid (TCA) cycle. Its expression can be transcriptionally regulated by p53 and Sp1, and its activity is modulated by upstream signals including HIF-1?? and mitochondrial stress. Downstream, ABAT controls levels of succinic semialdehyde, succinate, and glutamate, thereby influencing TCA cycle anaplerosis and NADH production. The enzyme physically interacts with ALDH5A1 (succinic semialdehyde dehydrogenase) and the mitochondrial import machinery to facilitate metabolic flux through this pathway.
In the context of HCT 116 colorectal carcinoma cells, ABAT knockout precipitates accumulation of GABA and a deficiency of succinate, disrupting the GABA shunt and impairing TCA cycle function. This metabolic lesion can compromise mitochondrial oxidative phosphorylation and reduce availability of biosynthetic precursors, potentially attenuating tumor cell proliferation and survival. The model is thus valuable for dissecting how colorectal tumors utilize GABA as a carbon source and for exploring metabolic vulnerabilities arising from ABAT deficiency, especially in the setting of KRAS-driven malignancies.
Researchers can employ these polyclonal knockout cells in a wide array of applications, including investigation of GABAergic metabolism in colorectal cancer, validation of ABAT as a therapeutic target, and screening of small-molecule ABAT inhibitors. Typical downstream assays encompass Western blotting and RT-qPCR for confirmation of ABAT disruption, HPLC- or ELISA-based GABA quantification, succinate level measurements, cell viability (MTT/CCK-8) and apoptosis (Annexin V/PI) analyses, Transwell migration/invasion assays, Seahorse metabolic flux analysis, and LC-MS-based TCA cycle metabolite profiling. These cells enable mechanistic studies on how the GABA shunt contributes to central carbon metabolism and tumor aggressiveness. For further technical details, please contact Ascent Research.
