Vav1 promotes pancreatic tumor cell invasion via GLS1-dependent glutamine metabolism

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy due in part to metabolic rewiring and a high rate of metastasis. Pancreatic tumors are particularly dependent upon glutamine (Gln) as a nutrient source for multiple metabolic pathways. To adapt to low levels of glutamine, PDAC cells upregulate the Rac1 dependent process of macropinocytosis, which internalizes extracellular macromolecules to scavenge them as a glutamine source. The proto oncogene Vav1 is an activator of the potent Rac/Cdc42 signaling cascades that regulate actin dynamics. While Vav1 is normally restricted to hematopoietic cells, Vav1 expression is significantly increased in PDAC, with high Vav1 correlating with worse overall survival. Here we uncover a novel Vav1 mediated regulation of glutamine metabolism in PDAC cells. While Vav1 is a potent activator of Rac1, surprisingly, Vav1 expression inhibited macropinocytosis in PDAC cells. Intriguingly, genomic and metabolomic analysis revealed that high Vav1 expression correlates with increased dependence on glutamine and glutamate (Glu) metabolism. Indeed, Vav1 dependent macropinocytosis was rescued by cell permeable glutamate, confirming disrupted Gln/Glu balance in Vav1 knockdown cells. RPPA screening indicated a Vav1 dependent effect on glutaminase 1 (GLS1, KGA isoform), which converts glutamine to glutamate. Mechanistically, we discovered that Vav1 modulates GLS1 localization to the mitochondria through regulation of GLS1 posttranslational modification. Disrupting GLS1 localization by deleting its mitochondrial targeting motif phenocopied loss of Vav1 in reducing the invasive potential of tumor cells. Investigation of Vav1 dependent metabolic consequences showed that knockdown of Vav1 alters 13C Gln derived metabolic flux into the TCA cycle, as well as a deficiency in the level of the antioxidant glutathione (GSH), which utilizes glutamate for its synthesis. Consequently, targeting Vav1 resulted in increased levels of cellular and mitochondrial reactive oxygen species (ROS) and increased sensitivity to lipid peroxidation. Notably, dual targeting of both glucose and glutamine pathways showed significant synergistic effects on cellular viability in Vav1 expressing cells. Here we established a new tumor promoting role for Vav1 via Gln/Glu regulation and uncover a novel subcellular regulation of mitochondrial glutamine metabolism. Our findings establish Vav1 as a critical regulator of metabolic rewiring in PDAC and orchestrates glutamine metabolism by controlling the KGA isoform of GLS1 mitochondrial localization. Importantly, Vav1 functions as a signaling adapter in glutamine metabolism, providing a potential therapeutic vulnerabilities for PDAC patients with high Vav1 expression.