A Role for the Krebs Cycle Intermediate Citrate in Metabolic Reprogramming in Innate Immunity and Inflammation

Abstract

Metabolism in immune cells is no longer thought of as merely a process for adenosine triphosphate (ATP) production, biosynthesis, and catabolism. The reprogramming of metabolic pathways upon activation is also for the production of metabolites that can act as immune signaling molecules. Activated dendritic cells (DCs) and macrophages have an altered Krebs cycle, one consequence of which is the accumulation of both citrate and succinate. Citrate is exported from the mitochondria via the mitochondrial citrate- carrier. Cytosolic metabolism of citrate to acetyl-coenzyme A (acetyl-CoA) is important for both fatty-acid synthesis and protein acetylation, both of which have been linked to macrophage and DC activation. Citrate-derived itaconate has a direct antibacterial effect and also has been shown to act as an anti-inflammatory agent, inhibiting succinate dehydrogenase. These findings identify citrate as an important metabolite for macrophage and DC effector function.

Keywords: ATP-citrate lyase; Krebs cycle; acetylation; citrate; immunometabolism; itaconate; macrophages; metabolism.

Figure 1

Citrate metabolism. Citrate is produced in the Krebs cycle from oxaloacetate and acetyl-CoA by citrate synthase (CS). It can be exported from the mitochondria through citrate carrier (CIC). Cytosolic citrate is broken down by ACLY to oxaloacetate and acetyl-CoA. Acetyl-CoA can be used as a substrate for fatty acid synthesis. High levels of cytosolic citrate can directly inhibit the glycolytic enzymes PFK1 and PFK2. PFK1 is also indirectly inhibited by decreased levels of fructose 2,6-bisphosphate and pyruvate kinase (PK) is indirectly inhibited (broken line) due to reduced levels of its activator, fructose-1,6-bisphosphate. Mitochondrial citrate can inhibit pyruvate dehydrogenase (PDH) and succinate dehydrogenase (SDH). Citrate-derived malonyl-CoA can block fatty acid oxidation (FAO) by inhibiting carnitine palmitoyltransferase 1 (CPT1). Citrate can activate the gluconeogenic enzyme fructose 1,6-bisphosphatase (FBPase1) and ACC, and so stimulate fatty acid synthesis.

Figure 2

Citrate in inflammation. Cytosolic citrate is broken down into acetyl-CoA and oxaloacetate. Further processing of oxaloacetate can provide a source of NADPH which is required by NAPDH oxidase and iNOS for the production of ROS and NO, respectively. Acetyl-CoA is required for acetylation of proteins and can be converted to malonyl-CoA for lysine-malonylation. Citrate-derived lipid synthesis has been linked to membrane expansion necessary for DC activation and cytokine production. In macrophages, the production of PGE₂ has been linked to citrate metabolism.

Figure 3

Citrate-derived itaconate. Citrate is converted to cis-aconitate by the Krebs cycle enzyme ACO2. Activation of macrophages with LPS causes induction of IRG1 which can produce itaconate by the decarboxylation of cis-aconitate. LPS also initiates the production of other pro-inflammatory mediators, such as HIF1 α, NO, and IL1β. Itaconate is toxic to microorganisms expressing ICL, a key component of the glycoxylate shunt in bacteria. Itaconate is also able to inhibit SDH and cause decreased production of NO, IL1β, IL18, and HIF1α.