BCAT1 inhibits glutamine-dependent Akt/mTOR signaling and nucleotide synthesis by initiating BTRC-mediated degradation of SLC3A2 in pancreatic adenocarcinoma

Pancreatic adenocarcinoma (PAAD) is characterized by profound metabolic reprogramming, including altered branched-chain amino acid (BCAA) metabolism. While the tumor-promoting role of branched-chain aminotransferase 2 (BCAT2) in PAAD has been well documented, the function of branched-chain aminotransferase 1 (BCAT1) remains unclear, particularly in PAAD cells with low endogenous BCAT1 expression. This study aimed to define the context-dependent role of BCAT1 in PAAD and to elucidate the underlying molecular mechanisms. The expression of BCAT1 and BCAA metabolism-related molecules was assessed by Western blotting and quantitative reverse transcription polymerase chain reaction (qRT-PCR). The effects of BCAT1 and SLC3A2 on PAAD cell proliferation were evaluated using CCK-8 assays in vitro and xenograft nude mouse models in vivo. Astral DIA proteomics and LC-MS-based untargeted metabolomics were performed to characterize protein and metabolic alterations induced by BCAT1 overexpression or SLC3A2 depletion. Stable SLC3A2-knockdown PAAD cell lines were established by lentiviral transduction. Seahorse XFe96 analysis was used to assess cellular bioenergetics by measuring the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR). BCAT1 expression was markedly reduced in multiple PAAD cell lines, and ectopic BCAT1 overexpression significantly inhibited PAAD cell proliferation both in vitro and in vivo. Mechanistically, BCAT1 overexpression suppressed Akt/mTOR signaling, impaired mitochondrial respiration and glycolytic activity, and disrupted glutamine metabolism, glutathione metabolism, and nucleotide biosynthesis. Notably, intracellular BCAA levels were not significantly altered, whereas glutamine was markedly reduced. Glutamine supplementation partially rescued the BCAT1-induced inhibition of Akt/mTOR signaling and cell growth. Proteomic analysis further revealed that BCAT1 overexpression reduced SLC3A2 expression, and SLC3A2 depletion phenocopied the effects of BCAT1 overexpression on glutamine metabolism, bioenergetics, Akt signaling, and tumor growth. Mechanistically, BCAT1 promoted SLC3A2 degradation through a BTRC-dependent proteasomal pathway, thereby restricting glutamine uptake and reducing intracellular nucleotide availability. BCAT1 functions as a context-dependent tumor suppressor in PAAD. In BCAT1-low PAAD cells, BCAT1 overexpression inhibits tumor growth by promoting BTRC-dependent proteasomal degradation of SLC3A2, thereby limiting glutamine uptake and suppressing Akt/mTOR signaling, cellular bioenergetics, and nucleotide biosynthesis, despite minimal changes in intracellular BCAA levels. These findings identify the BCAT1-BTRC-SLC3A2 axis as a previously unrecognized metabolic regulatory pathway in PAAD and suggest a potential therapeutic strategy for targeting tumor metabolic vulnerabilities.