Abstract

Idiopathic pulmonary fibrosis is characterized by the TGF-β-dependent differentiation of lung fibroblasts into myofibroblasts, leading to excessive deposition of extracellular matrix proteins, which distort lung architecture and function. Metabolic reprogramming in myofibroblasts is emerging as an important mechanism in the pathogenesis IPF and recent evidence suggests that glutamine metabolism is required in myofibroblasts although the exact role of glutamine in myofibroblasts is unclear. Here we demonstrate that glutamine, and its conversion to glutamate by glutaminase is required for TGF-β-induced collagen protein production in lung fibroblasts. We found that metabolism of glutamate to α-ketoglutarate by glutamate dehydrogenase or the glutamate-pyruvate or glutamate-oxaloacetate transaminases are not required for collagen protein production. Instead, we discovered that the glutamate-consuming enzymes phosphoserine aminotransferase 1 (PSAT1) and aldehyde dehydrogenase 18A1 (ALDH18A1)/ Δ1-pyrroline-5-carboxylate synthetase (P5CS) are required for collagen protein production by lung fibroblasts. PSAT1 is required for de novo glycine production while ALDH18A1/P5CS is required for de novo proline production. Consistent with this, we found that TGF-β treatment increased cellular levels of glycine and proline in lung fibroblasts. Our results suggest that glutamine metabolism is required to promote amino acid biosynthesis and not to provide intermediates such as α-ketoglutarate for oxidation in mitochondria. In support of this, we found that inhibition of glutaminolysis has no effect on cellular oxygen consumption and that knockdown of oxoglutarate dehydrogenase has no effect on the ability to produce collagen protein. Our results suggest that amino acid biosynthesis pathways may represent novel therapeutic targets for treatment of fibrotic diseases, including IPF.