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SLC15A4 mediates M1-prone metabolic shifts in macrophages and guards immune cells from metabolic stress

Significance SLC15A4 is a lysosome-resident amino acid/oligopeptide transporter that is preferentially expressed in immune cells. SLC15A4 is required for TLR7/9-dependent type I interferon production and plays a critical role in the pathogenesis of lupus and colitis. However, the precise molecular mechanisms of SLC15A4-mediated inflammatory responses are still undetermined. We revealed that SLC15A4 loss disturbed the coupling of glycolysis and the TCA cycle, and SLC15A4-deficient macrophages preferred to use glutamine rather than glucose as a carbon source for the TCA cycle. SLC15A4-deficient macrophages produced low levels of itaconate and proinflammatory IL-12 cytokine members. Our results reveal a mechanism of metabolic regulation in which an amino acid transporter acts as a guardian to allow macrophages to acquire an M1-prone metabolic phenotype. The amino acid and oligopeptide transporter Solute carrier family 15 member A4 (SLC15A4), which resides in lysosomes and is preferentially expressed in immune cells, plays critical roles in the pathogenesis of lupus and colitis in murine models. Toll-like receptor (TLR)7/9- and nucleotide-binding oligomerization domain-containing protein 1 (NOD1)-mediated inflammatory responses require SLC15A4 function for regulating the mechanistic target of rapamycin complex 1 (mTORC1) or transporting L-Ala-γ-D-Glu-meso-diaminopimelic acid, IL-12: interleukin-12 (Tri-DAP), respectively. Here, we further investigated the mechanism of how SLC15A4 directs inflammatory responses. Proximity-dependent biotin identification revealed glycolysis as highly enriched gene ontology terms. Fluxome analyses in macrophages indicated that SLC15A4 loss causes insufficient biotransformation of pyruvate to the tricarboxylic acid cycle, while increasing glutaminolysis to the cycle. Furthermore, SLC15A4 was required for M1-prone metabolic change and inflammatory IL-12 cytokine productions after TLR9 stimulation. SLC15A4 could be in close proximity to AMP-activated protein kinase (AMPK) and mTOR, and SLC15A4 deficiency impaired TLR-mediated AMPK activation. Interestingly, SLC15A4-intact but not SLC15A4-deficient macrophages became resistant to fluctuations in environmental nutrient levels by limiting the use of the glutamine source; thus, SLC15A4 was critical for macrophage’s respiratory homeostasis. Our findings reveal a mechanism of metabolic regulation in which an amino acid transporter acts as a gatekeeper that protects immune cells’ ability to acquire an M1-prone metabolic phenotype in inflammatory tissues by mitigating metabolic stress.