GPT2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population with targeted disruption of the GPT2 gene. This loss-of-function model enables investigation of mitochondrial alanine aminotransferase in a genetically simplified background, avoiding clonal selection bias. The polyclonal format captures population-level heterogeneity, ideal for functional genomics and metabolic studies.
The HAP1 cell line is a near-haploid chronic myeloid leukemia (CML) model, derived from a male patient, characterized by a predominantly haploid karyotype. This genetic simplicity allows unambiguous assignment of phenotypes to single gene disruptions, as only one allele requires modification for functional knockout. HAP1 cells maintain intact signaling pathways relevant to amino acid metabolism and cancer biology, providing a physiologically relevant context for studying GPT2.
GPT2 encodes mitochondrial alanine aminotransferase, which, using pyridoxal phosphate (PLP) as a cofactor, catalyzes reversible transamination of alanine and 2-oxoglutarate to pyruvate and glutamate. This reaction links amino acid metabolism to the TCA cycle and gluconeogenesis. GPT2 is regulated by PPARGC1A and glucagon and modulates downstream targets including GLUL, glutathione synthesis, and the pyruvate dehydrogenase complex. Its activity directly influences glutamate and alanine pools, intersecting with GLUD1-mediated deamination, thus integrating nitrogen and carbon metabolism critical for energy homeostasis and redox balance.
Disrupting GPT2 in HAP1 cells provides a powerful system for studying neurodevelopmental disorders such as intellectual disability and microcephaly, as well as hyperalaninemia. The near-haploid background amplifies metabolic shifts, enabling sensitive detection of changes in alanine, glutamate, and pyruvate. This model is particularly useful for dissecting gluconeogenic flux and nitrogen handling, processes often altered in cancer and metabolic diseases. Compensatory pathways involving alternative transaminases can also be investigated.
Applications include metabolic flux analysis via LC-MS metabolomics, alanine aminotransferase activity assays, and Seahorse metabolic profiling to assess mitochondrial function. The cells support drug target validation, amino acid dependency studies in cancer, and neurodevelopmental disease modeling. Standard assays such as Western blotting and RT-qPCR confirm GPT2 disruption and downstream effects. For further inquiries, contact Ascent Research.
