The GOT2 Knockout HeLa Polyclonal Cells comprise a CRISPR/Cas9-edited polyclonal population derived from HeLa cells, in which the GOT2 gene has been disrupted to create a loss-of-function model. This product provides a heterogeneous pool of GOT2-deficient HeLa cells, generated through targeted CRISPR/Cas9-mediated gene disruption, enabling functional investigation of mitochondrial aspartate aminotransferase without the selection of a single clone. The polyclonal format retains population-level diversity, making it suitable for studying gene essentiality and metabolic adaptations in a physiologically relevant cellular context.
The parental HeLa cell line is an immortalized human epithelial line originally derived from an HPV18-positive cervical adenocarcinoma. It is one of the most widely used models in cancer biology, cell cycle research, and cancer metabolism studies. HeLa cells exhibit robust proliferation, well-characterized signaling networks, and a high reliance on glutamine-driven anaplerosis, making them an ideal host for dissecting mitochondrial metabolic pathways. The combination of GOT2 knockout with this well-established cell system offers a powerful tool for investigating how mitochondrial transamination reactions sustain tumor cell growth and survival.
GOT2 encodes the mitochondrial isoform of aspartate aminotransferase, a pyridoxal phosphate-dependent enzyme that catalyzes the reversible transamination between aspartate and 2-oxoglutarate to yield oxaloacetate and glutamate. This reaction is central to the malate-aspartate shuttle, which transfers reducing equivalents from the cytosol into the mitochondria, maintaining NAD+/NADH balance and supporting oxidative phosphorylation. GOT2 is transcriptionally activated by ATF4 and MYC downstream of mTORC1 signaling and in response to amino acid deprivation or oxidative stress; NRF2 also regulates its expression under redox imbalance. The enzyme physically interacts with MDH2 and GOT1, and is deacetylated and activated by SIRT3. Its activity directly modulates intracellular aspartate pools, oxaloacetate availability, and glutamate homeostasis, thereby influencing nucleotide biosynthesis and TCA cycle anaplerosis. Key pathway partners include the mitochondrial carriers SLC25A11 (2-oxoglutarate/malate carrier) and SLC25A12 (aspartate/glutamate carrier), along with MDH1 and MDH2.
In the HeLa context, GOT2 disruption profoundly impacts cellular metabolism, as these cells depend on mitochondrial aspartate production for sustained nucleotide synthesis and NAD+ regeneration. Loss of GOT2 function impairs malate-aspartate shuttle activity, leading to cytosolic NADH accumulation, altered redox homeostasis, and reduced capacity for aerobic glycolysis. This model is therefore highly relevant for investigating metabolic vulnerabilities in cervical adenocarcinoma and other cancers, particularly the interplay between amino acid metabolism and cell proliferation. GOT2-deficient HeLa cells may exhibit enhanced sensitivity to inhibitors of glycolysis or mitochondrial respiration, providing a platform for identifying synthetic lethal interactions.
Researchers can employ these polyclonal knockout cells in a variety of experimental settings, including metabolic flux analysis using Seahorse analyzers, quantitative measurement of aspartate and glutamate by LC-MS, and assessment of NAD+/NADH ratios. Cell viability assays under glutamine limitation or electron transport chain inhibition can reveal metabolic dependencies. Immunofluorescence or Western blotting can validate GOT2 loss and monitor compensatory changes in GOT1 or MDH2. These cells are also suitable for RNA-seq or metabolomics to map global pathway remodeling. For further technical specifications and ordering information, please contact Ascent Research.
