Role of Polyamines in Immune Cell Functions

Abstract

The immune system is remarkably responsive to a myriad of invading microorganisms and provides continuous surveillance against tissue damage and developing tumor cells. To achieve these diverse functions, multiple soluble and cellular components must react in an orchestrated cascade of events to control the specificity, magnitude and persistence of the immune response. Numerous catabolic and anabolic processes are involved in this process, and prominent roles for l-arginine and l-glutamine catabolism have been described, as these amino acids serve as precursors of nitric oxide, creatine, agmatine, tricarboxylic acid cycle intermediates, nucleotides and other amino acids, as well as for ornithine, which is used to synthesize putrescine and the polyamines spermidine and spermine. Polyamines have several purported roles and high levels of polyamines are manifest in tumor cells as well in autoreactive B- and T-cells in autoimmune diseases. In the tumor microenvironment, l-arginine catabolism by both tumor cells and suppressive myeloid cells is known to dampen cytotoxic T-cell functions suggesting there might be links between polyamines and T-cell suppression. Here, we review studies suggesting roles of polyamines in normal immune cell function and highlight their connections to autoimmunity and anti-tumor immune cell function.

Keywords: B-lymphocytes; T-lymphocytes; autoimmunity; epigenetics; immunity; metabolism; tumor immunity.

Conflict of interest statement

The authors receive support from Celgene Corporation through a grant to the Moffitt Cancer Center & Research Institute. The content of this manuscript is not influenced by this association.

Figures

Figure 1

Stimulatory and inhibitory molecules expressed on T-cells. Diagram depicting the antigen presenting cell (APC) and T-cell interactions and activating receptors and ligands on these cells that govern the functional outcomes of T-cells such as cytotoxicity-associated granzyme B expression, cytokine release, and proliferation [61]. The T-cell receptor complex is composed of several proteins that are necessary for survival and signaling including T-cell receptor (TCR)α and TCRβ chains, CD3 signaling molecules δ/ε, CD3γ/ε and CD247 composed on the dimeric ζζ-chains or ζn (not shown). Co-stimulatory molecules on T-cells such as CD28, the founding member of the immunoglobulin (Ig) family of costimulatory receptors, are critical to amplify and sustain the signaling response. Activation leads to metabolic reprogramming to increase glycolysis, oxidative phosphorylation, and amino acid metabolism through glutaminolysis and ultimately to polyamine biosynthesis [62,63]. Additional receptors include CD40L (CD154), T-cell specific surface glycoprotein CD28, inducible T-cell costimulatory ICOS (CD278) which is a CD28-family molecule expressed on T-cells important for Th2 responses, Traf-linked tumor necrosis factor receptor family protein, CD27, which is important in T and B-cell memory formation and activation of natural killer (NK) cells [64,65,66], tumor necrosis factor receptor superfamily (TNFRSF), member 4 (TNFRSF4) also OX40 (CD134) expressed on activated T-cells [51,52,53], leukocyte-associated antigen-1 (LFA1) which is an integrin involved in T-cell migration [53,67], the adhesion molecule CD2 present on T-cells and NK cells (also known erythrocyte receptor and rosette receptor, LFA-2), herpesvirus entry mediator (HVEM) also known as tumor necrosis factor receptor superfamily member 14 (TNFRSF14), glucocorticoid-induced TNFR family related gene (GITR) a member of the TNFRSF [68], S-type lectin Galectin 9, T-cell immunoglobulin mucin domain 1 (TIM1) also known as hepatitis A virus cellular receptor 1 (HAVcr-1), and 4-1BB (CD137, TNFRS9). Corresponding receptors on APC are the classical costimulatory ligands CD80 (B7-1), CD86 (B7-2) that interact with CD28, TNFRS5 (CD40), human inducible costimulatory-ligand (ICOSL) [69], ligand for CD27 (CD70 also TNFSF7), OX40 ligand (OX40L), intercellular adhesion molecule 1 (ICAM-1, also CD54), leukocyte-associated antigen-3 (LFA3), HVEM counter-receptor lymphotoxin-like, exhibits inducible expression, and competes with herpes simplex virus glycoprotein D for HVEM, a receptor expressed by T-lymphocytes (LIGHT, also CD160), GITR ligand (GITRL), and 4-1BB ligand (4-1BBL). Inhibitory receptors and ligands are shown including cytotoxic T-lymphocyte antigen-4 (CTLA4) which interacts with CD80, CD86 that also recognizes CD28, programmed cell death protein 1 (PD1) receptor and its ligands PD-ligand 1 (PDL1) and PD-ligand 2 (PDL2), and B-and T-lymphocyte attenuator (BTLA) and CD160 [70] that both recognize HVEM. MHC: major histocompatibility complex, Ag: antigenic peptide.

Figure 2

Proximal T-cell signaling cascade. Proximal signaling pathways downstream of the T-cell receptor (TCR)-antigen presenting cell (APC)ignaling complex (as described in Figure 1) are responsible for the cascade of events leading to metabolic reprogramming including the transcription of amino acid transporter and enzymes involved in metabolism of nutrients and biosynthesis of polyamines [85]. Phosphorylation of the immunoreceptor tyrosine-based activation motifs (ITAMs) on the cytoplasmic side of the TCR/CD3 complex engage numerous cascading interactions largely mediated by phosphorylation, dephosphorylation or ubiquitinylation resulting in cellular activation [61]. The initiating signal is generated by lymphocyte protein tyrosine kinase (Lck) and other proto-oncogene tyrosine-protein kinase (Src) family tyrosine kinases including the zeta-chain associated protein kinase (Zap-70) that is recruited to the TCR/CD3 complex. Costimulation through leukocyte-associated antigen-1 (LFA1) which is an integrin involved in T-cell migration or CD28 interaction with CD80 (B7-1) or CD86 (B7-2) (see also Figure 1) activates the phosphorylation of the YXXM or YNPP signaling motifs [86] which regulates glucose metabolism. CD28 leads to stable recruitment of the adaptor protein Grb2/GADS along with interleukin-2-indicible T-cell kinase (Itk), Lck, and phosphatidylinositide 3 kinase (PI3K) heterodimer p85/p110 and SLP76. These interactions promote the activation of VAV-1, RasGRP, and the Ras/Raf/MEK/Erk pathway downstream of phosphorylated SLP-76 and Zap-70 modulating the TCR signal strength [86]. A complement of transcription factors nuclear factor of activated T-cells (NFAT), cAMP response element-binding protein (CREB), Fox family transcription factor c-Fos, Jun (when in combination with c-Fos forms the AP-1 early response transcription factor complex, nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), an NFκB family member c-Rel, and c-Myc which coordinately regulate gene expression. Activation of CD28 leads to the phosphorylation of PI3K, phosphatidylinositol-3,4 bisphosphate (PIP2) and phosphoinositide-dependent kinase 1 (PDK1) [87] which integrates the TCR and CD28 signaling to induce the NFκB pathway including protein kinase C-theta (PKC-θ), and inhibits the ubiquitin ligase c-Cbl [88] leading to activation of Bcl10, Malt1, Carma1 (CBM) complex leading to IKKαβγ activation of NFκB and REL [87]. In addition to PKC-θ, phosphorylation of Akt is critical for the regulation of mTORC1 and mTORC2 complexes of mTOR that bind GβL and raptor or rictor, respectively [79,81]. This is a critical step in c-Myc-dependent transcriptional regulation that stimulates dramatic changes in metabolism including glucose, amino acid, nucleotide and polyamine biosynthesis [63,89]. Divalent cations such as calcium (Ca2+) are induce downstream of phospholipase C γ1, PIP2, and indo inositol-1,4,5 triphosphate (IP3) which mobilizes the release of intracellular Ca2+ stores from the endoplasmic reticulum (Ca2+-ER) a potential metabolic switch that suppresses intratumoral T-cell function [90]. Sustained signaling then promotes the influx of extracellular Ca2+ into the cells through calcium release-activated Ca2+ (CRAC) channels. Calcium-calmodulin interactions (Ca2+/CaM) then activates the phosphatase calcineurin and calcium/calmodulin-dependent protein kinase type IV calmodulin (CaMKIV), which dephosphorylates the cytoplasmic subunits of nuclear factor of activated T-cells (NFAT) exposing a nuclear localization signal resulting in nuclear transport and phosphorylates CREB, respectively.

Figure 3

Bioenergetics of macrophage subsets. Monocyte-derived macrophages can be differentially polarized by the cytokine milieu [125,126]. (A) M1 macrophages originate from cells in the bone marrow and develop in inflammatory environments. Nitric oxide (NO) is the major byproduct of these cells arising from the reaction of arginine with oxygen through the actions of inducible nitric oxide synthase (iNOS) which produces citrulline and NO (see detailed pathway Figure 3C). Citrulline is then exported and re-imported to re-generate arginine and sustain NO production. A product of the degradation of arginine through this cycle is fumarate which is derived from the conversion of argininosuccinate to arginine (see Figure 3C). M1 macrophages are also critical for the production of cytokines and chemokines and for the production of itaconate which acts as an anti-microbial cellular metabolite. Succinate, a proinflammatory molecule that controls IL-1β expression, accumulates and stabilizes the oxygen sensing pathway regulated by hyposia-inducible factor 1-alpha (HIF1α) [125,127]. (B) Unlike M1 macrophages, the polarized M2 subtype reduces their ability to make NO and instead hydrolyzes imported arginine into ornithine and urea through the urea cycle (detailes in Figure 3C). M2 macrophages are therefore suppressive by competing for both arginine and glutamine that is necessary for effector T-cell functions [63,89,112]. To fuel their functions, including proliferation, M2 macrophages use fatty acids oxidation (FAO) which supports oxidative phosphorylation and electron transport through the tricarboxylic acid (TCA) cycle. Also present in the suppressive tumor microenvironment is a population of bone marrow derived immature myeloid cells known as myeloid derived suppressor cells [128,129]. While bioenergetics for these cells needs further analysis, they retain NO production and FAO, TCA and deplete arginine and glutamine [130] from the microenvironment. (C) Also detailed is the metabolism of arginine, l-citrulline and l-ornithine to produce fumarate from conversion of argininosuccinate. Citrulline plus aspartate generates argininosuccinate via the actions of argininosuccinate synthetase (ASS) in the cytosol and ornithine is converted to citrulline by carbamylphosphate plus ornithine via the enzymatic activity of ornithine transcarbamoylase (OTC). Additional enzymes and reactions include those metabolized by ODC: ornithinine decarboxylase, ARG: arginase 1 or arginase 2, ADC: arginine decarboxylase which is the biosynthetic enzyme for agmatate [131], OAT: ornithine aminotransferase, NOS: nitric oxide synthase, PRMT: protein arginine methyltransferases which is important for epigenetic regulation [132], and AGAT: l-arginine:glycine amidinotransferase which is the enzyme that catalyzes the transfer of an amidino group from l-arginine to produce l-ornithine and guanidinoacetate and acts as the immediate precursor of creatine.

Figure 3

Bioenergetics of macrophage subsets. Monocyte-derived macrophages can be differentially polarized by the cytokine milieu [125,126]. (A) M1 macrophages originate from cells in the bone marrow and develop in inflammatory environments. Nitric oxide (NO) is the major byproduct of these cells arising from the reaction of arginine with oxygen through the actions of inducible nitric oxide synthase (iNOS) which produces citrulline and NO (see detailed pathway Figure 3C). Citrulline is then exported and re-imported to re-generate arginine and sustain NO production. A product of the degradation of arginine through this cycle is fumarate which is derived from the conversion of argininosuccinate to arginine (see Figure 3C). M1 macrophages are also critical for the production of cytokines and chemokines and for the production of itaconate which acts as an anti-microbial cellular metabolite. Succinate, a proinflammatory molecule that controls IL-1β expression, accumulates and stabilizes the oxygen sensing pathway regulated by hyposia-inducible factor 1-alpha (HIF1α) [125,127]. (B) Unlike M1 macrophages, the polarized M2 subtype reduces their ability to make NO and instead hydrolyzes imported arginine into ornithine and urea through the urea cycle (detailes in Figure 3C). M2 macrophages are therefore suppressive by competing for both arginine and glutamine that is necessary for effector T-cell functions [63,89,112]. To fuel their functions, including proliferation, M2 macrophages use fatty acids oxidation (FAO) which supports oxidative phosphorylation and electron transport through the tricarboxylic acid (TCA) cycle. Also present in the suppressive tumor microenvironment is a population of bone marrow derived immature myeloid cells known as myeloid derived suppressor cells [128,129]. While bioenergetics for these cells needs further analysis, they retain NO production and FAO, TCA and deplete arginine and glutamine [130] from the microenvironment. (C) Also detailed is the metabolism of arginine, l-citrulline and l-ornithine to produce fumarate from conversion of argininosuccinate. Citrulline plus aspartate generates argininosuccinate via the actions of argininosuccinate synthetase (ASS) in the cytosol and ornithine is converted to citrulline by carbamylphosphate plus ornithine via the enzymatic activity of ornithine transcarbamoylase (OTC). Additional enzymes and reactions include those metabolized by ODC: ornithinine decarboxylase, ARG: arginase 1 or arginase 2, ADC: arginine decarboxylase which is the biosynthetic enzyme for agmatate [131], OAT: ornithine aminotransferase, NOS: nitric oxide synthase, PRMT: protein arginine methyltransferases which is important for epigenetic regulation [132], and AGAT: l-arginine:glycine amidinotransferase which is the enzyme that catalyzes the transfer of an amidino group from l-arginine to produce l-ornithine and guanidinoacetate and acts as the immediate precursor of creatine.