Hu Zeng

mTORC1 couples immune signals and metabolic programming to establish T reg -cell function

The mechanistic target of rapamycin (mTOR) pathway integrates diverse environmental inputs, including immune signals and metabolic cues, to direct T-cell fate decisions. The activation of mTOR, which is the catalytic subunit of the mTORC1 and mTORC2 complexes, delivers an obligatory signal for the proper activation and differentiation of effector CD4+ T cells, whereas in the regulatory T-cell (Treg) compartment, the Akt–mTOR axis is widely acknowledged as a crucial negative regulator of Treg-cell de novo differentiation and population expansion. However, whether mTOR signalling affects the homeostasis and function of Treg cells remains largely unexplored. Here we show that mTORC1 signalling is a pivotal positive determinant of Treg-cell function in mice. Treg cells have elevated steady-state mTORC1 activity compared to naive T cells. Signals through the T-cell antigen receptor (TCR) and interleukin-2 (IL-2) provide major inputs for mTORC1 activation, which in turn programs the suppressive function of Treg cells. Disruption of mTORC1 through Treg-specific deletion of the essential component raptor leads to a profound loss of Treg-cell suppressive activity in vivo and the development of a fatal early onset inflammatory disorder. Mechanistically, raptor/mTORC1 signalling in Treg cells promotes cholesterol and lipid metabolism, with the mevalonate pathway particularly important for coordinating Treg-cell proliferation and upregulation of the suppressive molecules CTLA4 and ICOS to establish Treg-cell functional competency. By contrast, mTORC1 does not directly affect the expression of Foxp3 or anti- and pro-inflammatory cytokines in Treg cells, suggesting a non-conventional mechanism for Treg-cell functional regulation. Finally, we provide evidence that mTORC1 maintains Treg-cell function partly through inhibiting the mTORC2 pathway. Our results demonstrate that mTORC1 acts as a fundamental ‘rheostat’ in Treg cells to link immunological signals from TCR and IL-2 to lipogenic pathways and functional fitness, and highlight a central role of metabolic programming of Treg-cell suppressive activity in immune homeostasis and tolerance.

T Cell Exit from Quiescence and Differentiation into Th2 Cells Depend on Raptor-mTORC1-Mediated Metabolic Reprogramming

Naive T cells respond to antigen stimulation by exiting from quiescence and initiating clonal expansion and functional differentiation, but the control mechanism is elusive. Here we describe that Raptor-mTORC1-dependent metabolic reprogramming is a central determinant of this transitional process. Loss of Raptor abrogated T cell priming and T helper 2 (Th2) cell differentiation, although Raptor function is less important for continuous proliferation of actively cycling cells. mTORC1 coordinated multiple metabolic programs in T cells including glycolysis, lipid synthesis, and oxidative phosphorylation to mediate antigen-triggered exit from quiescence. mTORC1 further linked glucose metabolism to the initiation of Th2 cell differentiation by orchestrating cytokine receptor expression and cytokine responsiveness. Activation of Raptor-mTORC1 integrated T cell receptor and CD28 costimulatory signals in antigen-stimulated T cells. Our studies identify a Raptor-mTORC1-dependent pathway linking signal-dependent metabolic reprogramming to quiescence exit, and this in turn coordinates lymphocyte activation and fate decisions in adaptive immunity.

Bcl10 plays a critical role in NF-κB activation induced by G protein-coupled receptors

G protein-coupled receptors (GPCRs) play pivotal roles in cell proliferation, differentiation, and survival. Although many studies indicate that the stimulation of GPCRs leads to NF-κB activation, the molecular mechanism by which GPCRs induced NF-κB activation remains largely unknown. Bcl10 is an essential adaptor molecule connecting antigen receptor signaling cascades to NF-κB activation in lymphocytes. However, the function of Bcl10 in nonlymphoid cells remains to be determined. In this study, we demonstrated that the deficiency of Bcl10 resulted in the defect in NF-κB activation induced by either expressing the constitutively active mutant of G protein or stimulation of cells with lysophosphatidic acid or endothelin-1, which activate their GPCR. In contrast, TNF-α-, LPS-, and integrin-induced NF-κB activation was not affected in Bcl10-deficient cells. Together, our results provide genetic evidence showing that Bcl10 is a key signaling component mediating NF-κB activation induced by GPCRs in nonlymphoid cells.

Tuberous sclerosis 1 (Tsc1)-dependent metabolic checkpoint controls development of dendritic cells

Significance A fundamental question in immunology is how the coordination of immune signals and metabolic programs regulates immune responses. The identification of metabolic pathways orchestrating the activation of lymphocytes and dendritic cells (DCs) has advanced our understanding of immune activation, but whether cell metabolism contributes to development of immune cells is unknown. Here we have genetically defined a crucial metabolic checkpoint for DC development that is mediated by the interplay between Tsc1-mTOR complex 1 signaling and Myc-dependent bioenergetic and biosynthetic programs. Dysregulation of this pathway impairs survival, proliferation, and functional differentiation of DCs, thereby highlighting the importance of metabolic programming of DC development. Coordination of cell metabolism and immune signals is crucial for lymphocyte priming. Emerging evidence also highlights the importance of cell metabolism for the activation of innate immunity upon pathogen challenge, but there is little evidence of how this process contributes to immune cell development. Here we show that differentiation of dendritic cells (DCs) from bone marrow precursors is associated with dynamic regulation of mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) signaling and cell metabolism. Unexpectedly, enhancing mTORC1 activity via ablation of its negative regulator tuberous sclerosis 1 (Tsc1) impaired DC development in vivo and in vitro, associated with defective cell survival and proliferation. Moreover, Tsc1 deficiency caused DC spontaneous maturation but a propensity to differentiate into other lineages, and attenuated DC-mediated effector TH1 responses. Mechanistically, Tsc1-deficient DCs exhibited increased glycolysis, mitochondrial respiration, and lipid synthesis that were partly mediated by the transcription factor Myc, highlighting a key role of Tsc1 in modulating metabolic programming of DC differentiation. Further, Tsc1 signaled through Rheb to down-regulate mTORC1 for proper DC development, whereas its effect at modulating mTOR complex 2 (mTORC2) activity was largely dispensable. Our results demonstrate that the interplay between Tsc1-Rheb-mTORC1 signaling and Myc-dependent bioenergetic and biosynthetic activities constitutes a key metabolic checkpoint to orchestrate DC development.

Bcl10 Plays a Divergent Role in NK Cell-Mediated Cytotoxicity and Cytokine Generation

Activating receptors such as NKG2D and Ly49D mediate a multitude of effector functions including cytotoxicity and cytokine generation in NK cells. However, specific signaling events that are responsible for the divergence of distinct effector functions have yet to be determined. In this study, we show that lack of caspase recruitment domain-containing protein Bcl10 significantly affected receptor-mediated cytokine and chemokine generation, but not cytotoxicity against tumor cells representing “missing-self” or “induced-self.” Lack of Bcl10 completely abrogated the generation of GM-CSF and chemokines and it significantly reduced the generation of IFN-γ (>75%) in NK cells. Commitment, development, and terminal maturation of NK cells were largely unaffected in the absence of Bcl10. Although IL-2-activated NK cells could mediate cytotoxicity to the full extent, the ability of the freshly isolated NK cells to mediate cytotoxicity was somewhat reduced. Therefore, we conclude that the Carma1-Bcl10-Malt1 signaling axis is critical for cytokine and chemokine generation, although it is dispensable for cytotoxic granule release depending on the activation state of NK cells. These results indicate that Bcl10 represents an exclusive “molecular switch” that links the upstream receptor-mediated signaling to cytokine and chemokine generations.

Phosphorylation of Bcl10 Negatively Regulates T-Cell Receptor-Mediated NF-κB Activation

ABSTRACT Bcl10 (B-cell lymphoma 10) is an adaptor protein comprised of an N-terminal caspase recruitment domain and a C-terminal serine/threonine-rich domain. Bcl10 plays a critical role in antigen receptor-mediated NF-κB activation and lymphocyte development and functions. Our current study has discovered that T-cell activation induced monophosphorylation and biphosphorylation of Bcl10 and has identified S138 within Bcl10 as one of the T-cell receptor-induced phosphorylation sites. Alteration of S138 to an alanine residue impaired T-cell activation-induced ubiquitination and subsequent degradation of Bcl10, ultimately resulting in prolongation of TCR-mediated NF-κB activation and enhancement of interleukin-2 production. Taken together, our findings demonstrate that phosphorylation of Bcl10 at S138 down-regulates Bcl10 protein levels and thus negatively regulates T-cell receptor-mediated NF-κB activation.

Essential Role of Phospholipase Cγ2 in Early B-Cell Development and Myc-Mediated Lymphomagenesis

ABSTRACT Phospholipase Cγ2 (PLCγ2) is a critical signaling effector of the B-cell receptor (BCR). Here we show that PLCγ2 deficiency impedes early B-cell development, resulting in an increase of B220+ CD43+ BP-1+ CD24hi pre-BCR+ large pre-B cells. PLCγ2 deficiency impairs pre-BCR-mediated functions, leading to enhanced interleukin-7 (IL-7) signaling and elevated levels of RAGs in the selected large pre-B cells. Consequently, PLCγ2 deficiency renders large pre-B cells susceptible to transformation, resulting in dramatic acceleration of Myc-induced lymphomagenesis. PLCγ2−/− Eμ-Myc transgenic mice mainly develop lymphomas of B220+ CD43+ BP-1+ CD24hi pre-BCR+ large pre-B-cell origin, which are uncommon in wild-type Eμ-Myc transgenics. Furthermore, lymphomas from PLCγ2−/− Eμ-Myc transgenic mice exhibited a loss of p27Kip1 and often displayed alterations in Arf or p53. Thus, PLCγ2 plays an important role in pre-BCR-mediated early B-cell development, and its deficiency leads to markedly increased pools of the most at-risk large pre-B cells, which display hyperresponsiveness to IL-7 and express high levels of RAGs, making them prone to secondary mutations and Myc-induced malignancy.

B Cell Lymphoma 10 Is Essential for FcεR-Mediated Degranulation and IL-6 Production in Mast Cells

The adaptor protein B cell lymphoma 10 (Bcl10) plays an essential role in the functions of the AgRs in T and B cells. In this study, we report that Bcl10 also plays an important role in mast cells. Bcl10 is expressed in mast cells. Although Bcl10-deficient mast cells undergo normal development, we demonstrate that Bcl10 is essential for specific functions of FcεR. Although Bcl10-deficient mast cells have normal de novo synthesis and release of the lipid mediator arachidonic acid, the mutant cells possess impaired FcεR-mediated degranulation, indicated by decreased serotonin release, and impaired cytokine production, measured by release of IL-6. In addition, Bcl10-deficient mice display impaired IgE-mediated passive cutaneous anaphylaxis. Moreover, although Bcl10-deficient mast cells have normal FcεR-mediated Ca2+ flux, activation of PI3K, and activation of the three types of MAPKs (ERKs, JNK, and p38), the mutant cells have markedly diminished FcεR-mediated activation of NF-κB and decreased activation of AP-1. Thus, Bcl10 is essential for FcεR-induced activation of AP-1, NF-κB, degranulation, and cytokine production in mast cells.

mTOR and metabolic regulation of conventional and regulatory T cells

mTOR signaling links bioenergetic and biosynthetic metabolism to immune responses. mTOR is activated by diverse upstream stimuli, including immune signals, growth factors, and nutrients. Recent studies highlight crucial roles of mTOR signaling in immune functions mediated by conventional T cells and Tregs. In this review, we discuss the regulation of mTOR signaling in T cells and the functional impacts of mTOR and metabolic pathways on T cell‐mediated immune responses, with a particular focus on the differentiation and function of Tregs.

T Cell Receptor-mediated Activation of CD4+CD44hi T Cells Bypasses Bcl10 AN IMPLICATION OF DIFFERENTIAL NF-κB DEPENDENCE OF NAïVE AND MEMORY T CELLS DURING T CELL RECEPTOR-MEDIATED RESPONSES

Previous studies have demonstrated that Bcl10 (B-cell leukemia/lymphoma 10) is essential for T cell receptor-mediated NF-κB activation and subsequent proliferation and interleukin 2 (IL2) production. However, here we demonstrate that, contrary to expectations, Bcl10 is differentially required for T cell activation, including for both proliferation and cytokine production. When CD4+ and CD8+ T cells were divided based on expression levels of CD44, which distinguishes naïve cells (CD44lo) versus those that are antigen-experienced (CD44hi), IL2 production by and proliferation of CD4+CD44lo naïve cells and both subpopulations of CD8+ T cells were clearly Bcl10-dependent, whereas these same functional properties of CD4+CD44hi T cells occurred largely independent of Bcl10. As with the other subpopulations of T cells, CD4+CD44hi T cells did not activate the NF-κB pathway in the absence of Bcl10; nevertheless, these CD4+CD44hi antigen-experienced T cells efficiently secreted IL2 after T cell receptor stimulation. Strikingly, therefore, T cell receptor-mediated IL2 production in these cells is NF-κB-independent. Our studies suggest that antigen-experienced CD4+ T cells differ from their naïve counterparts and from CD8+ T cells in their ability to achieve activation independent of the Bcl10/NF-κB pathway.