Regulation of immune responses by mTOR

Abstract

mTOR is an evolutionarily conserved serine/threonine kinase that plays a central role in integrating environmental cues in the form of growth factors, amino acids, and energy. In the study of the immune system, mTOR is emerging as a critical regulator of immune function because of its role in sensing and integrating cues from the immune microenvironment. With the greater appreciation of cellular metabolism as an important regulator of immune cell function, mTOR is proving to be a vital link between immune function and metabolism. In this review, we discuss the ability of mTOR to direct the adaptive immune response. Specifically, we focus on the role of mTOR in promoting differentiation, activation, and function in T cells, B cells, and antigen-presenting cells.

Figure 1

mTOR integrates environmental cues to direct the adaptive immune response. Antigen-presenting cells (APCs), T cells, and B cells play central roles in the adaptive immune response. This figure highlights the diverse environmental cues integrated by mTOR, which are discussed in this review.

Figure 2

Structure of the mTOR signaling complex. (a) mTOR is an evolutionarily conserved 289-kDa serine/threonine protein kinase that is composed of two N-terminal HEAT (huntingtin, elongation factor 3, subunit of PP2A, and TOR) domains, which mediate protein-protein interactions, adjacent to a FRAP, ATM, and TRRAP (FAT) domain. The FRB domain is where the small 12-kDa FK506-binding protein (FKBP12) bound to the macrolide drug rapamycin binds to mTOR to inhibit its activity. The mTOR kinase catalytic domain lies C-terminal to the FRB site, and this is where mTOR kinase inhibitors bind. The carboxy FAT (FATC) domain maintains the structural integrity of this large protein kinase. (b) mTOR associates with two distinct sets of adapter proteins to form two intracellular signaling complexes with unique substrate specificity. The TORC1 signaling complex is composed of the regulatory-associated protein of mTOR (RAPTOR) and mammalian lethal with Sec13 protein 8 (mLST8), which are both adapter proteins that mediate protein-protein interactions via their WD-40 domains. The proline-rich Akt substrate 40 kDa (PRAS40) and DEP domain–containing mTOR-interacting protein (DEPTOR) inhibit mTORC1 activity. The mTORC2 complex can also associate with DEPTOR and mLST8, but this complex is distinguished by the adapter protein RAPTOR-independent companion of TOR (RICTOR) and protein observed with RICTOR (PROTOR). Another unique component of mTORC2 is mSIN1, which contains a pleckstrin homology domain that is thought to target TORC2 to the membrane, where it can activate myristoylated Akt.

Figure 3

The upstream signaling cascade leading to mTOR activation in an immune setting. Growth factor stimulation leads to the recruitment of PI3 kinase (PI3K), which phosphorylates phosphatidylinositol 4,5-bisphosphate (PIP2) at the 3′ position to generate phosphatidylinositol 3,4,5-trisphosphate (PIP3). This leads to the recruitment of Akt to the membrane to be phosphorylated (at position T308) by PDK1. Activated Akt phosphorylates TSC2 in an inhibitory manner, yielding a separation of the TSC1/TSC2 complex and loss of GAP activity for RHEB-GTP. This leads to an accumulation of RHEB-GTP, which promotes mTORC1 function. Similarly, Erk can phosphorylate TSC2 in an inhibitory manner. Alternatively, the following phosphorylate TSC2 in an activating way, thereby enhancing its GAP activity: AMPK, in response to low levels of energy; REDD1, in response to low oxygen tension; and GSK3β, which is regulated by WNT and Akt. In the presence of amino acids, Rag proteins bind to Raptor and promote the relocalization of mTORC1 with Rheb-GTP, leading to activation. Rapamycin bound to FKBP12 allosterically inhibits mTORC1 activity. Additionally, many immunologic inputs also play a role in regulating mTORC1 activity. Positive costimulation (such as CD28 engagement) as well as cytokine signaling lead to recruitment of PI3K activity. Conversely, PD-1 ligation (co-inhibition) inhibits PI3K function. The upstream regulation of mTORC2 is poorly understood. Growth factor stimulation activates this complex, whereas high doses or prolonged exposure to rapamycin will disrupt the mTORC2 complex. Dashed lines indicate that the exact mechanism is unknown. Black lines show activating signals, red lines show inhibitory signals, and red arrows indicate signals that indirectly lead to inhibition of mTORC1 activity.

Figure 4

Downstream mTORC1 and mTORC2 signaling. Upon activation of mTORC1, mTOR phosphorylates S6K1, leading to the phosphorylation of ribosomal S6 protein, which allows for enhanced protein translation. Phosphorylation of 4E-BP1 by mTOR releases eIF-4E to participate in the translation-initiation complexes. Along with increasing protein translation, mTORC1 activity also upregulates gene expression programs necessary for glucose and lipid metabolism, mitochondrial biogenesis, and inhibition of autophagy. Immunologically, mTORC1 activity leads to the inhibition of SOCS3 and the increased activation of STAT4 and STAT3. This in turn leads to increases in T-bet and ROR γt in response to IL-12 and IL-6, respectively, which promote Th1 and Th17 differentiation. mTORC2 activity leads to the phosphorylation of Akt (at position S473) and SGK1, leading to their activation and in turn resulting in the phosphorylation and sequestration of FOXO proteins in the cytoplasm. This prevents the FOXO proteins from activating the transcription of target genes such as Krüppel-like factor 2 (KLF2), which itself influences the expression of CD62L, CCR7, and S1P1. mTORC2 activity also inhibits SOCS5 expression, thereby enhancing STAT6 phosphorylation in response to IL-4 and subsequent GATA-3 expression and Th2 differentiation. In addition, mTORC2 signaling activates PKCθ, which in turn can also promote Th2 differentiation. Dashed lines indicate that the exact mechanism is unknown, black lines show activating signals, and red lines show inhibitory signals.

Figure 5

Pharmacologic inhibitors of mTOR and calcineurin signaling. Rapamycin, cyclosporin A (CsA), and FK506 are all immunosuppressive agents with similar structure yet diverse mechanisms of action. All three agents are activated by binding to cis-trans peptidyl-prolyl isomerase proteins called immunophilins. CsA binds to cyclophilin, whereas FK506 and rapamycin bind to FKBP12. Both CsA and FK506 inhibit TCR-induced calcineurin activation and thus inhibit the translocation of NF-AT to the nucleus. In doing so, they block the expression of NF-AT-dependent genes of activation such as IL-2 as well as NF-AT-induced inhibitory genes such as Cbl-b. As such, in addition to being potent immunosuppressive agents, these compounds also block the induction of TCR-induced tolerance. However, rapamycin and similar drugs termed rapalogs, when bound to FKBP12, inhibit the interaction of RAPTOR and mTOR and thus inhibit mTORC1 activation. Prolonged exposure of T cells to rapamycin can also impair mTORC2 activity by an as yet undefined mechanism. In contrast, mTOR kinase inhibitors function as ATP-competitive inhibitors at the mTOR catalytic domain to specifically and potently inhibit both mTORC1 and mTORC2. Wortmannin and LY94002 can block mTOR activity by inhibiting PI3K activity, which is upstream of mTOR. Additionally, molecules that mimic the effects of AMP, such as metformin, can inhibit mTOR activity by activating AMPK, which in turn promotes the ability of TSC2 to inhibit mTORC1. Dashed lines indicate that the exact mechanism is unknown, black lines show activating signals, and red lines show inhibitory signals.

Figure 6

The role of mTOR in CD4⁺ T cell differentiation. A series of genetic studies has revealed a central role for mTOR in regulating T helper cell differentiation. T cells lacking RHEB, and hence mTORC1 signaling, fail to differentiate into Th1 and Th17 cells under polarizing conditions. This failure to differentiate is associated with decreased STAT activation and decreased expression of lineage-specific transcription factors such as T-bet and RORγt. Similarly, T cells lacking RICTOR and mTORC2 activity demonstrate decreased STAT6 activation, decreased PKC activity, and decreased GATA-3 expression and thus fail to differentiate into Th2 cells under polarizing conditions. T cells lacking mTOR demonstrate increased SMAD3 activation and become Foxp3⁺ Tregs even under normally activating conditions. Black lines indicate the ability of mTOR to activate a pathway, and red lines indicate the ability of mTOR to inhibit a pathway.

Figure 7

mTOR regulates maturation and function of antigen-presenting cells. Plasmacytoid dendritic cells (DCs) activated by TLR9 and TLR7 agonists in the presence of rapamycin fail to produce interferon-α and -β, indicating an important role for mTOR in promoting type 1 interferon expression (a). Activation of monocytes and DCs with LPS in the presence of rapamycin leads to an increase in IL-12 production and a decrease in IL-10 production. Such findings suggest that mTOR inhibits proinflammatory gene expression in these cells (c). Alternatively, rapamycin blocks the maturation of bone marrow–derived DCs (b). Such cells display decreased MHC and costimulatory molecules and actually promote the induction of anergic and regulatory T cells. Interestingly, in spite of the ability of the rapamycin-matured DCs to promote tolerance, they also produce increased IL-12 upon stimulation with LPS.