BCAT2 Knockout Jurkat Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population that disrupts the branched-chain amino acid transaminase 2 (BCAT2) gene in the Jurkat human T-lymphocyte cell line. This loss-of-function model enables the study of BCAT2-dependent branched-chain amino acid (BCAA) metabolism and its impact on T-cell signaling and leukemogenesis.
The Jurkat cell line, derived from the peripheral blood of a patient with acute T-cell leukemia, is a well-established model for investigating T-cell receptor signaling, activation, and leukemia biology. These immortalized T lymphocytes retain key signaling pathways and metabolic programs relevant to primary T cells, making them a valuable platform for gene-editing studies.
BCAT2 encodes the mitochondrial isoform of branched-chain amino acid aminotransferase, which catalyzes the reversible transamination of leucine, isoleucine, and valine to their corresponding ??-keto acids????-ketoisocaproate, ??-keto-??-methylvalerate, and ??-ketoisovalerate??using pyridoxal phosphate as a cofactor. This reaction is the first step in BCAA catabolism and provides carbon skeletons for TCA cycle anaplerosis and energy production. BCAT2 expression is regulated transcriptionally by c-MYC and HIF1A, and its activity influences the cellular pool of glutamate and ??-keto acids. Downstream, BCAT2 contributes to mTORC1 activation via branched-chain keto acid dehydrogenase (BCKD) complex?Cdependent production of acetyl-CoA and succinyl-CoA, as well as through modulation of intracellular amino acid levels sensed by mTORC1 signaling complexes. The BCKD complex comprises BCKDHA, BCKDHB, DBT, and DLD subunits.
In Jurkat cells, BCAT2 disruption abrogates BCAA transamination, leading to accumulation of BCAAs and depletion of ??-keto acids, which disrupts anaplerotic influx into the TCA cycle. This metabolic perturbation impairs mTORC1 signaling, as indicated by reduced phosphorylation of downstream effectors such as ribosomal protein S6 kinase (S6K), and compromises T-cell activation programs, including CD69 upregulation and IL-2 secretion. Consequently, this polyclonal knockout population serves as a physiologically relevant system to dissect how BCAA catabolism couples metabolic fitness to T-cell functional responses and leukemic growth.
Researchers can employ this knockout tool to explore metabolic vulnerabilities in T-cell leukemia, examine the role of BCAT2 in cancer metabolic reprogramming, and investigate how BCAA availability modulates mTORC1 signaling in activated T cells. Representative assays include Western blotting and RT-qPCR for confirming BCAT2 loss, intracellular BCAA quantification, 13C-BCAA metabolic tracing to track carbon flux, phospho-S6K flow cytometry, CD69 and IL-2 ELISA for activation phenotypes, and Seahorse metabolic flux analysis to assess oxidative and glycolytic activity. For technical inquiries or ordering information, please contact Ascent Research.
