Metabolic Regulation of Natural Killer Cell IFN-γ Production

Abstract

Metabolism is critical for a host of cellular functions and provides a source of intracellular energy. It has been recognized recently that metabolism also regulates differentiation and effector functions of immune cells. Although initial work in this field has focused largely on T lymphocytes, recent studies have demonstrated metabolic control of innate immune cells, including natural killer (NK) cells. Here, we review what is known regarding the metabolic requirements for NK cell activation, focusing on NK cell production of interferon-gamma (IFN-γ). NK cells are innate immune lymphocytes that are poised for rapid activation during the early immune response. Although their basal metabolic rates do not change with short-term activation, they exhibit specific metabolic requirements for activation depending upon the stimulus received. These metabolic requirements for NK cell activation are altered by culturing NK cells with interleukin-15, which increases NK cell metabolic rates at baseline and shifts them toward aerobic glycolysis. We discuss the metabolic pathways important for NK cell production of IFN-γ protein and potential mechanisms whereby metabolism regulates NK cell function.

FIG. 1

Signaling pathways leading to IFN-γ transcription in NK cells. NK cells up-regulate the transcription of Ifng in response to several signaling pathways, most of which converge on the TFs STAT4 and NF-κB to cause acute transcription. In NK cells, the Ifng locus is bound by constitutively active T-bet and Eomes. Shown here are the primary signaling pathways downstream of cytokines and receptors leading to IFN-γ transcription. IL-12-induced STAT4 is essential for optimal cytokine co-stimulation of IFN-γ. IL-2 and IL-15 share common signaling receptors and downstream Janus kinase (JAK)/STAT, PI3K, and MAPK signaling. There is evidence that NF-κB and STAT4 can also be activated downstream of IL-2 in NK cells, although this signaling is poorly described (lightened). Activation receptors can trigger IFN-γ production independently of cytokine signaling and associate with ITAM-containing adapters, leading to multiple downstream signaling cascades including PI3K, MAPK, and PLC-γ, which cause cytokine production and cytotoxicity. Red indicates ligand; blue, receptor; green, kinase; purple, transcription factor; and teal, second messenger.

FIG. 2

Major metabolic pathways and programs in immune cells. Left: Naive immune cells use glucose to fuel glycolysis, producing ATP and pyruvate. Pyruvate is converted into acetyl-CoA, enters into the mitochondria, and is processed by the TCA cycle into high-energy electrons. Carriers bring these electrons to the electron transport chain for OXPHOS, which requires oxygen to synthesize large amounts of ATP. Center: In activated effector T cells, DCs, and M1 macrophages, glycolysis is highly up-regulated, leading to the production of lactate. Metabolites are shunted toward nucleotide synthesis in T cells, ROS/nitric oxide in macrophages, and fatty acid synthesis in DCs. Depleted TCA intermediates can be replaced by glutamine. OXPHOS functions at a low level in activated effector cells. Right: Alternatively activated M2 macrophages, Tregs, and memory T cells down-regulate glycolysis, instead oxidizing lipids to fuel the TCA cycle. Increased mitochondrial mass leads to greater respiratory capacity and increased OXPHOS.

FIG. 3

Increased rates of metabolism with prolonged IL-15 exposure, but not short-term activation. Short-term activation (6 hours) by NKRs or cytokines does not change the rates of glycolysis or OXPHOS despite the production of IFN-γ and degranulation. Culture with IL-15 for 3+ days increases both glycolysis and OXPHOS, with a greater enhancement of glycolysis but, by itself, does not lead to significant IFN-γ production. These IL-15-“primed” NK cells also have different metabolic requirements for IFN-γ production compared with naive NK cells.

FIG. 4

Naive NK cells require OXPHOS for receptor-stimulated IFN-γ production, but activation with IL-15 eliminates this requirement. Fresh murine splenic NK cells (top and middle) or IL-15-activated “primed” NK cells (bottom, 72-hour stimulation with 100 ng/mL IL-15) were activated with IL-12/18 (top) or plate-bound anti-NK1.1 (center and bottom) in the presence or absence of the OXPHOS inhibitor oligomycin (100 nM). NK cell production of IFN-γ protein was measured after 6 hours.