ACTIVATED T‐CELLS UNDERGO AN ESSENTIAL METABOLIC SHIFT
Metabolic reprogramming drives T‐cell activation and effector differentiation
It should now be apparent that lymphocyte activation triggers a myriad of signaling pathways that radically transform resting T‐cells in preparation for effector function, and recent developments have uncovered a crucial role for specific metabolic pathways in not only fueling these changes, but in directing the outcome of T‐cell differentiation into specific effector subtypes. Activated T‐cells not only differ metabolically from their quiescent counterparts, differentiation into the various effector populations cannot proceed without distinct metabolic reprogramming.
Naive T‐cells are constantly on the move, migrating through lymphoid tissues to patrol for signs of infection by endlessly sampling MHC–peptide complexes displayed by antigen‐presenting cells. This dynamism, driven by constant cytoskeletal reorganization, is extremely energy demanding and requires an efficient method of ATP generation while at the same time necessitating minimal new biosynthesis. Resting T‐cells use the highly efficient ATP‐generating process of oxidative phosphorylation (OXPHOS) to supply their energy requirements. In simple terms, glucose is first broken down to pyruvate in the cytoplasm in a separate process called glycolysis, which also generates 2 molecules of ATP (Figure 7.16). Pyruvate is then converted into acetyl‐CoA, the apical factor in a series of chemical reactions in the mitochondria called the tricarboxylic acid (TCA) cycle. The endpoint of the TCA cycle is the generate NADH for use as an electron donor in the mitochondrial electron transport chain, the oxygen‐dependent process of OXPHOS, which produces up to 34 additional molecules of ATP from a single glucose molecule. In addition, fatty acids and certain amino acids can be catabolized to supply acetyl‐CoA to drive the TCA cycle and supply the electron transport chain. Therefore, resting T‐cells use the OXPHOS pathway to convert the majority of their nutrient supply, in the form of sugar, fatty acids, and protein, into ATP.
Figure 7.16 Metabolic pathways driving growth and proliferation. Glycolysis and the tricarboxylic acid (TCA) cycle function separately and in combination to generate ATP and biosynthesis‐promoting metabolites. Glucose is first broken down into pyruvate, which can then be converted to NAD+ and used to re‐start glycolysis. A small amount of pyruvate can also be used as a source of acetyl‐CoA to drive the TCA cycle in mitochondria. Intermediates from the glycolysis pathway can be siphoned off and used by the pentose phosphate pathway to produce ribose‐5P and by the serine biosynthesis pathway to generate serine, both of which can be used to make nucleotides. Citrate can be removed from the TCA cycle and used to regenerate acetyl‐CoA for lipid biosynthesis. To keep the TCA cycle moving in the absence of citrate, glutamine is converted to glutamate through glutaminolysis, and then to α‐ketoglutarate, to re‐enter the cycle. Oxaloacetate can also be used to generate aspartate for nucleotide synthesis.
In low oxygen conditions, cells can make do with the less efficient glycolysis pathway to generate their energy needs, where instead of being used as an intermediate to drive the TCA cycle, glucose‐derived pyruvate is converted to lactate to generate NAD+ to restart the glycolytic process, yielding a meagre 2 molecules of ATP per molecule of glucose (Figure 7.16). It may come as a surprise then, that activated T‐cells primarily use glycolysis to generate ATP even in the presence of oxygen, in a process termed aerobic glycolysis. While counterintuitive at first, an appreciation of what T‐cell activation sets out to achieve sheds light on this strange choice of energy‐producing system. In contrast to quiescent T‐cells, which require abundant energy but only minimal levels of new biosynthesis, activated T‐cells must rapidly proliferate and differentiate into effector T‐cells to meet the challenge of infection, a process that not only requires energy but also considerable new biosynthesis to generate daughter cells and inflammatory cytokines. While generating a low yield of ATP, glycolysis produces a wealth of metabolites essential for building new cells and proteins. Importantly, derivatives from glycolysis can be siphoned off and used in the pentose phosphate and serine biosynthesis pathways to produce an abundant supply of metabolic precursors for nucleotide and fatty acid synthesis (Figure 7.16). During aerobic glycolysis, most pyruvate is converted to lactate but a small amount is shunted into a modified TCA cycle, where citrate is extracted and used to synthesize lipids crucial for building cell membranes, while fatty acid catabolism is actively inhibited. To keep this improvised TCA cycle moving, glutamine is converted to glutamate and then to α‐ketoglutarate in a process called glutaminolysis, which replaces the α‐ketoglutarate that would otherwise have been generated by citrate. In summary, activated T‐cells turn the majority of their nutrient supply into biomass to build an army of antigen‐specific daughter cells and crucial inflammatory mediators with which to fight an infection (Figure 7.16).
Signals from the TCR, co‐stimulatory molecules, and cytokines coordinate the metabolism of activated T‐cells
Although many cell types tailor their metabolic profile based on the nutrients available to them, the transition of antigenstimulated T‐cells to aerobic glycolysis is driven by signals propagated directly by the TCR complex, maintained by CD28 co‐stimulation, and fine‐tuned by inflammatory cytokines. TCR stimulation directly controls the switch from OXPHOS, where nutrients are consumed to generate ATP, to a glycolytic, biomass‐generating metabolism to support the biosynthesis needed for daughter cells and inflammatory mediators. TCR triggering transmits these signals through serine‐threonine kinases, which turn on a range of transcription factors and crucial regulators that together coordinate an increase in glucose and amino acid uptake essential for driving glycolysis while at the same time, blocking the oxidation of lipids to favor fatty acid synthesis to build cell membranes.
The transcription factor c‐Myc is activated early on TCR
stimulation by RAS‐activated ERK1 and ERK2, and plays a crucial role in regulating glycolysis. c‐Myc promotes the expression of essential glycolytic genes, including the cell membrane glucose transporter Glut1; glutaminase, which drives glutaminolysis; lactate dehydrogenase, essential for converting pyruvate into lactate to replenish the glycolytic cycle; and a number of glutamine transporters (Figure 7.16). The importance of glycolysis for T‐cell activation is illustrated by the fate of c‐Myc‐deficient T‐cells, which are completely incompetent for glycolysis and glutaminolysis and fail to proliferate on antigen stimulation.
TCR triggering also induces the expression of l‐amino acid transporters on the cell membrane, promoting the influx of leucine, critical for activation of another important glycolysis regulator, the mTORC1 complex. Increased intracellular leucine shunts mTORC1 to the lysosomal membrane, where it can be activated by RAS homolog enriched in brain (RHEB). Importantly, activation of the CD28 co‐receptor complex is required for mTORC1 activation as CD28‐induced activation of PI3K, in collaboration with mTORC2, facilitates AKT‐ mediated inactivation of the mTORC1 repressor TSC2, leading to mTORC1 activation. mTORC1 has multiple effects on T‐cell activation, by increasing the rate of protein translation and by blocking fatty acid oxidation through SREBP2‐mediated inhibition of CPT1a, a protein required for supplying the mitochondria with fatty acids to burn during OXPHOS. mTORC1 also activates hypoxia‐inducible factor 1α (HIF1α), a transcription factor well known for promoting the expression of genes required for survival in oxygen‐deprived environments. HIF1α induces many of the genes required for glycolysis, and while the mTORC1/HIF1α axis is not required for activating glycolysis at the early stages of TCR stimulation, their activation is essential for promoting the sustained glycolysis required for full T‐cell activation. As CD28 triggering also activates glutamine trans porters required for glutaminolysis, co‐receptor stimulation through mTORC1/HIF1α activation and glutamine import plays a crucial role in sustaining T‐cell activation long enough for proliferation to occur, a point further illustrated by the action of the immunosupressant drug rapamycin, which directly inhibits the mTORC1 complex, leading to a state of T‐cell anergy. Thus, the coordinated efforts of c‐Myc, the mTOR complex and HIF1α trigger a switch to a glycolytic metabolism essential for fueling biosynthesis of antigen‐specific daughter cells and inflammatory cytokines required to fight infection.
Conversely, activation of the glycolysis pathway is opposed by the cytosolic nutrient sensor AMPK, which blocks the activation of mTORC1, inhibiting glycolysis, and facilitating the accumulation of CPT1a, which promotes lipid oxidation in mitochondria, thus blocking lipid biosynthesis. AMPK is activated by an increased ratio of AMP:ATP, which indicates a drop in cellular ATP levels. In a finely tuned system of regulation, sufficient cellular ATP levels allow ATP to bind to and block an activating phosphorylation site on AMPK, preventing its phosphorylation by the AMPK‐activating kinase LBK1. When the cellular energy level drops and the AMP:ATP ratio increases, AMP can displace ATP from binding AMPK, thereby facilitating its activation by LBK1, and the promotion of XPHOS and ATP generation over glycolysis and biosynthesis.