RESUMO
Cell identity and function largely rely on the programming of transcriptomes during development and differentiation. Signature gene expression programs are orchestrated by regulatory circuits consisting of cis-acting promoters and enhancers, which respond to a plethora of cues via the action of transcription factors. In turn, transcription factors direct epigenetic modifications to revise chromatin landscapes, and drive contacts between distal promoter-enhancer combinations. In immune cells, regulatory circuits for effector genes are especially complex and flexible, utilizing distinct sets of transcription factors and enhancers, depending on the cues each cell type receives during an infection, after sensing cellular damage, or upon encountering a tumor. Here, we review major players in the coordination of gene regulatory programs within innate and adaptive immune cells, as well as integrative omics approaches that can be leveraged to decipher their underlying circuitry.
Assuntos
Cromatina , Redes Reguladoras de Genes , Animais , Regulação da Expressão Gênica , Humanos , Regiões Promotoras Genéticas , Fatores de Transcrição/genéticaRESUMO
Intestinal intraepithelial lymphocytes (IELs) exhibit prompt innate-like responses to microenvironmental cues and require strict control of effector functions. Here we showed that Aiolos, an Ikaros zinc-finger family member encoded by Ikzf3, acted as a regulator of IEL activation. Ikzf3-/- CD8αα+ IELs had elevated expression of NK receptors, cytotoxic enzymes, cytokines and chemokines. Single-cell RNA sequencing of Ikzf3-/- and Ikzf3+/+ IELs showed an amplified effector machinery in Ikzf3-/- CD8αα+ IELs compared to Ikzf3+/+ counterparts. Ikzf3-/- CD8αα+ IELs had increased responsiveness to interleukin-15, which explained a substantial part, but not all, of the observed phenotypes. Aiolos binding sites were close to those for the transcription factors STAT5 and RUNX, which promote interleukin-15 signaling and cytolytic programs, and Ikzf3 deficiency partially increased chromatin accessibility and histone acetylation in these regions. Ikzf3 deficiency in mice enhanced susceptibility to colitis, underscoring the relevance of Aiolos in regulating the effector function in IELs.
Assuntos
Linfócitos Intraepiteliais , Fatores de Transcrição , Animais , Camundongos , Antígenos CD8/metabolismo , Interleucina-15/metabolismo , Mucosa Intestinal/metabolismo , Linfócitos Intraepiteliais/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Innate lymphocytes encompass a diverse array of phenotypic identities with specialized functions. DNA methylation and hydroxymethylation are essential for epigenetic fidelity and fate commitment. The landscapes of these modifications are unknown in innate lymphocytes. Here, we characterized the whole-genome distribution of methyl-CpG and 5-hydroxymethylcytosine (5hmC) in mouse innate lymphoid cell 3 (ILC3), ILC2 and natural killer (NK) cells. We identified differentially methylated regions (DMRs) and differentially hydroxymethylated regions (DHMRs) between ILC and NK cell subsets and correlated them with transcriptional signatures. We associated lineage-determining transcription factors (LDTFs) with demethylation and demonstrated unique patterns of DNA methylation/hydroxymethylation in relationship to open chromatin regions (OCRs), histone modifications and TF-binding sites. We further identified an association between hydroxymethylation and NK cell superenhancers (SEs). Using mice lacking the DNA hydroxymethylase TET2, we showed the requirement for TET2 in optimal production of hallmark cytokines by ILC3s and interleukin-17A (IL-17A) by inflammatory ILC2s. These findings provide a powerful resource for studying innate lymphocyte epigenetic regulation and decode the regulatory logic governing their identity.
Assuntos
Metilação de DNA , Imunidade Inata , Animais , Cromatina/genética , Epigênese Genética , Imunidade Inata/genética , Células Matadoras Naturais , Linfócitos , CamundongosRESUMO
Natural killer (NK) cells develop from common progenitors but diverge into distinct subsets, which differ in cytokine production, cytotoxicity, homing, and memory traits. Given their promise in adoptive cell therapies for cancer, a deeper understanding of regulatory modules controlling clinically beneficial NK phenotypes is of high priority. We report integrated "-omics" analysis of human NK subsets, which revealed super-enhancers associated with gene cohorts that may coordinate NK functions and localization. A transcription factor-based regulatory scheme also emerged, which is evolutionarily conserved and shared by innate and adaptive lymphocytes. For both NK and T lineages, a TCF1-LEF1-MYC axis dominated the regulatory landscape of long-lived, proliferative subsets that traffic to lymph nodes. In contrast, effector populations circulating between blood and peripheral tissues shared a PRDM1-dominant landscape. This resource defines transcriptional modules, regulated by feedback loops, which may be leveraged to enhance phenotypes for NK cell-based therapies.
Assuntos
Células Matadoras Naturais/classificação , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/fisiologia , Citocinas/imunologia , Citocinas/metabolismo , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Humanos , FenótipoRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
Innate lymphoid cells (ILCs) are tissue-resident lymphocytes categorized on the basis of their core regulatory programs and the expression of signature cytokines. Human ILC3s that produce the cytokine interleukin-22 convert into ILC1-like cells that produce interferon-γ in vitro, but whether this conversion occurs in vivo remains unclear. In the present study we found that ILC3s and ILC1s in human tonsils represented the ends of a spectrum that included additional discrete subsets. RNA velocity analysis identified an intermediate ILC3-ILC1 cluster, which had strong directionality toward ILC1s. In humanized mice, the acquisition of ILC1 features by ILC3s showed tissue dependency. Chromatin studies indicated that the transcription factors Aiolos and T-bet cooperated to repress regulatory elements active in ILC3s. A transitional ILC3-ILC1 population was also detected in the human intestine. We conclude that ILC3s undergo conversion into ILC1-like cells in human tissues in vivo, and that tissue factors and Aiolos were required for this process.
Assuntos
Imunidade Inata/imunologia , Interferon gama/metabolismo , Interleucinas/metabolismo , Mucosa Intestinal/imunologia , Linfócitos/imunologia , Tonsila Palatina/imunologia , Animais , Diferenciação Celular/imunologia , Células Cultivadas , Criança , Pré-Escolar , Humanos , Fator de Transcrição Ikaros/metabolismo , Mucosa Intestinal/citologia , Linfócitos/classificação , Linfócitos/citologia , Camundongos , Proteínas com Domínio T/metabolismo , Interleucina 22RESUMO
Innate lymphoid cells (ILCs) serve as sentinels in mucosal tissues, sensing release of soluble inflammatory mediators, rapidly communicating danger via cytokine secretion, and functioning as guardians of tissue homeostasis. Although ILCs have been extensively studied in model organisms, little is known about these "first responders" in humans, especially their lineage and functional kinships to cytokine-secreting T helper (Th) cell counterparts. Here, we report gene regulatory circuitries for four human ILC-Th counterparts derived from mucosal environments, revealing that each ILC subset diverges as a distinct lineage from Th and circulating natural killer cells but shares circuitry devoted to functional polarization with their Th counterparts. Super-enhancers demarcate cohorts of cell-identity genes in each lineage, uncovering new modes of regulation for signature cytokines, new molecules that likely impart important functions to ILCs, and potential mechanisms for autoimmune disease SNP associations within ILC-Th subsets.
Assuntos
Linfócitos/citologia , Linfócitos/imunologia , Imunidade Adaptativa , Animais , Citocinas/imunologia , Citocinas/metabolismo , Elementos Facilitadores Genéticos , Humanos , Imunidade Inata , Imunidade nas Mucosas , Células Matadoras Naturais , Linfócitos/metabolismo , Camundongos , Tonsila Palatina/citologia , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Fatores de Transcrição/metabolismo , TranscriptomaRESUMO
In the mammalian intestine, crypts of Leiberkühn house intestinal epithelial stem/progenitor cells at their base. The mammalian intestine also harbors a diverse array of microbial metabolite compounds that potentially modulate stem/progenitor cell activity. Unbiased screening identified butyrate, a prominent bacterial metabolite, as a potent inhibitor of intestinal stem/progenitor proliferation at physiologic concentrations. During homeostasis, differentiated colonocytes metabolized butyrate likely preventing it from reaching proliferating epithelial stem/progenitor cells within the crypt. Exposure of stem/progenitor cells in vivo to butyrate through either mucosal injury or application to a naturally crypt-less host organism led to inhibition of proliferation and delayed wound repair. The mechanism of butyrate action depended on the transcription factor Foxo3. Our findings indicate that mammalian crypt architecture protects stem/progenitor cell proliferation in part through a metabolic barrier formed by differentiated colonocytes that consume butyrate and stimulate future studies on the interplay of host anatomy and microbiome metabolism.
Assuntos
Bactérias/metabolismo , Butiratos/metabolismo , Colo/citologia , Colo/microbiologia , Microbioma Gastrointestinal , Células-Tronco/metabolismo , Acil-CoA Desidrogenase/deficiência , Acil-CoA Desidrogenase/genética , Animais , Proliferação de Células , Intestino Delgado/citologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Oxirredução , Moléculas com Motivos Associados a Patógenos/metabolismo , Células-Tronco/citologia , Peixe-ZebraRESUMO
In contrast to the "helper" activities of most CD4+ T effector subsets, CD4+ cytotoxic T lymphocytes (CD4-CTLs) perform functions normally associated with CD8+ T and NK cells. Specifically, CD4-CTLs secrete cytotoxic molecules and directly target and kill compromised cells in an MHC class II-restricted fashion. The functions of these cells have been described in diverse immunological contexts, including their ability to provide protection during antiviral and antitumor responses, as well as being implicated in autoimmunity. Despite their significance to human health, the complete mechanisms that govern their programming remain unclear. In this article, we identify the Ikaros zinc finger transcription factor Eos (Ikzf4) as a positive regulator of CD4-CTL differentiation during murine immune responses against influenza virus infection. We find that the frequency of Eos+ cells is elevated in lung CD4-CTL populations and that the cytotoxic gene program is compromised in Eos-deficient CD4+ T cells. Consequently, we observe a reduced frequency and number of lung-residing, influenza virus-responsive CD4-CTLs in the absence of Eos. Mechanistically, we determine that this is due, at least in part, to reduced expression of IL-2 and IL-15 cytokine receptor subunits on the surface of Eos-deficient CD4+ T cells, both of which support the CD4-CTL program. Finally, we find that Aiolos, a related Ikaros family member and known CD4-CTL antagonist, represses Eos expression by antagonizing STAT5-dependent activation of the Ikzf4 promoter. Collectively, our findings reveal a mechanism wherein Eos and Aiolos act in opposition to regulate cytotoxic programming of CD4+ T cells.
Assuntos
Antineoplásicos , Linfócitos T CD4-Positivos , Camundongos , Humanos , Animais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Linfócitos T Citotóxicos , Diferenciação Celular , Citocinas/metabolismo , Antineoplásicos/metabolismoRESUMO
In the periphery, T helper cell differentiation is a key event orchestrating the adaptive immune response yet recent studies demonstrate considerable plasticity in these cell fate decisions. In this issue of Immunity, Mukasa et al. (2010) describe the epigenetic basis underlying this plasticity.
RESUMO
Follicular (FO) and marginal zone (MZ) B cells are maintained in distinct locations within the spleen, but the genetic basis for this separation is still enigmatic. We now report that B cell sequestration requires lineage-specific regulation of migratory receptors by the transcription factor Klf2. Moreover, using gene-targeted mice we show that altered splenic B cell migration confers a significant in vivo gain-of-function phenotype to FO B cells, including the ability to quickly respond to MZ-associated antigens and pathogens in a T cell-dependent manner. This work demonstrates that in wild-type animals, naive FO B cells are actively removed from the MZ, thus restricting their capacity to respond to blood-borne pathogens.
Assuntos
Linfócitos B/citologia , Linfócitos B/imunologia , Movimento Celular , Imunidade Humoral , Baço/citologia , Baço/imunologia , Animais , Antígenos CD19/genética , Antígenos CD19/imunologia , Antígenos T-Independentes/genética , Antígenos T-Independentes/imunologia , Medula Óssea/imunologia , Diferenciação Celular , Células Cultivadas , Fatores de Transcrição Kruppel-Like/deficiência , Fatores de Transcrição Kruppel-Like/imunologia , Camundongos , Camundongos Knockout , Receptores CCR/imunologiaRESUMO
Genome stability relies on epigenetic mechanisms that enforce repression of endogenous retroviruses (ERVs). Current evidence suggests that distinct chromatin-based mechanisms repress ERVs in cells of embryonic origin (histone methylation dominant) vs. more differentiated cells (DNA methylation dominant). However, the latter aspect of this model has not been tested. Remarkably, and in contrast to the prevailing model, we find that repressive histone methylation catalyzed by the enzyme SETDB1 is critical for suppression of specific ERV families and exogenous retroviruses in committed B-lineage cells from adult mice. The profile of ERV activation in SETDB1-deficient B cells is distinct from that observed in corresponding embryonic tissues, despite the loss of repressive chromatin modifications at all ERVs. We provide evidence that, on loss of SETDB1, ERVs are activated in a lineage-specific manner depending on the set of transcription factors available to target proviral regulatory elements. These findings have important implications for genome stability in somatic cells, as well as the interface between epigenetic repression and viral latency.
Assuntos
Linfócitos B/metabolismo , Retrovirus Endógenos/genética , Histona-Lisina N-Metiltransferase/genética , Retroviridae/genética , Animais , Linfócitos B/virologia , Western Blotting , Células da Medula Óssea/metabolismo , Células da Medula Óssea/virologia , Células Cultivadas , Metilação de DNA , Retrovirus Endógenos/metabolismo , Epigênese Genética , Citometria de Fluxo , Perfilação da Expressão Gênica , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Interações Hospedeiro-Patógeno , Lisina/metabolismo , Metilação , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Camundongos Transgênicos , Análise de Sequência com Séries de Oligonucleotídeos , Células Precursoras de Linfócitos B/metabolismo , Células Precursoras de Linfócitos B/virologia , Retroviridae/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sequências Repetidas Terminais/genética , Ativação Viral/genéticaRESUMO
Regulatory T cells (Tregs) are a specialized subset of CD4(+) T cells that maintain self-tolerance by functionally suppressing autoreactive lymphocytes. The Treg compartment is composed of thymus-derived Tregs (tTregs) and peripheral Tregs (pTregs) that are generated in secondary lymphoid organs after exposure to antigen and specific cytokines, such as TGF-ß. With regard to this latter lineage, pTregs [and their ex vivo generated counterparts, induced Tregs (iTregs)] offer particular therapeutic potential because these cells can be raised against specific antigens to limit autoimmunity. We now report that transcription factor Krüppel-like factor 2 (KLF2) is necessary for the generation of iTregs but not tTregs. Moreover, drugs that limit KLF2 proteolysis during T-cell activation enhance iTreg development. To the authors' knowledge, this study identifies the first transcription factor to distinguish between i/pTreg and tTreg ontogeny and demonstrates that KLF2 is a therapeutic target for the production of regulatory T cells.
Assuntos
Autoimunidade/imunologia , Diferenciação Celular/imunologia , Fatores de Transcrição Kruppel-Like/metabolismo , Linfócitos T Reguladores/imunologia , Animais , Autoimunidade/genética , Imunoprecipitação da Cromatina , Primers do DNA/genética , Citometria de Fluxo , Fatores de Transcrição Kruppel-Like/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos TransgênicosRESUMO
Previous studies have identified multiple conserved noncoding sequences (CNS) at the mouse Ifng locus sufficient for enhancer activity in cell-based assays. These studies do not directly address biology of the human IFNG locus in a genomic setting. IFNG enhancers may be functionally redundant or each may be functionally unique. We test the hypothesis that each IFNG enhancer has a unique necessary function using a bacterial artificial chromosome transgenic model. We find that CNS-30, CNS-4, and CNS+20 are required at distinct stages of Th1 differentiation, whereas CNS-16 has a repressive role in Th1 and Th2 cells. CNS+20 is required for IFN-γ expression by memory Th1 cells and NKT cells. CNS-4 is required for IFN-γ expression by effector Th1 cells. In contrast, CNS-16, CNS-4, and CNS+20 are each partially required for human IFN-γ expression by NK cells. Thus, IFNG CNS enhancers have redundant necessary functions in NK cells but unique necessary functions in Th cells. These results also demonstrate that distinct CNSs are required to transcribe IFNG at each stage of the Th1 differentiation pathway.
Assuntos
Interferon gama/genética , RNA não Traduzido/genética , Sequências Reguladoras de Ácido Nucleico , Células Th1/metabolismo , Animais , Diferenciação Celular/genética , Células Cultivadas , Cromossomos Artificiais Bacterianos , Sequência Conservada , Regulação da Expressão Gênica , Humanos , Interferon gama/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células T Matadoras Naturais/citologia , Células T Matadoras Naturais/imunologia , Células T Matadoras Naturais/metabolismo , Regiões Promotoras Genéticas , Células Th1/citologia , Células Th1/imunologia , Células Th2/citologia , Células Th2/imunologia , Células Th2/metabolismoRESUMO
The majority of the genome is noncoding and was thought to be nonfunctional. However, it is now appreciated that transcriptional control of protein coding genes resides within these noncoding regions. Thousands of genes encoding long intergenic noncoding RNAs (lincRNAs) have been recently identified throughout the genome, which positively or negatively regulate transcription of neighboring target genes. Both TMEVPG1 and its mouse ortholog encode lincRNAs and are positioned near the IFN-γ gene (IFNG). In this study, we show that transcription of both mouse and human TMEVPG1 genes is Th1 selective and dependent on Stat4 and T-bet, transcription factors that drive the Th1 differentiation program. Ifng expression is partially restored in Stat4-/-Tbx21-/- cells through coexpression of T-bet and Tmevpg1, and Tmevpg1 expression contributes to, but alone is not sufficient to, drive Th1-dependent Ifng expression. Our results suggest that TMEVPG1 belongs to the general class of lincRNAs that positively regulate gene transcription.
Assuntos
Regulação Viral da Expressão Gênica/imunologia , Interferon gama/genética , RNA não Traduzido/imunologia , Células Th1/imunologia , Células Th1/virologia , Theilovirus/genética , Theilovirus/imunologia , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Células Cultivadas , Humanos , Interferon gama/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Transgênicos , RNA Viral/genética , RNA Viral/imunologia , Células Th1/metabolismo , Regulação para Cima/genética , Regulação para Cima/imunologiaRESUMO
Antibody-dependent cell-mediated cytotoxicity, mediated by natural killer (NK) cells and antibodies, emerged as a secondary correlate of protection in the RV144 HIV vaccine clinical trial, the only vaccine thus far demonstrating some efficacy in human. Therefore, leveraging NK cells with enhanced cytotoxic effector responses may bolster vaccine induced protection against HIV. Here, we investigated the effect of orally administering indole-3-carbinol (I3C), an aryl hydrocarbon receptor (AHR) agonist, as an adjuvant to an RV144-like vaccine platform in a mouse model. We demonstrate the expansion of KLRG1-expressing NK cells induced by the vaccine together with I3C. This NK cell subset exhibited enhanced vaccine antigen-specific cytotoxic memory-like features. Our study underscores the potential of incorporating I3C as an oral adjuvant to HIV vaccine platforms to enhance antigen-specific (memory-like) cytotoxicity of NK cells against HIV-infected cells. This approach may contribute to enhancing the protective efficacy of HIV preventive vaccines against HIV acquisition.
RESUMO
Cutaneous leishmaniasis caused by Leishmania parasites exhibits a wide range of clinical manifestations. Although parasites influence disease severity, cytolytic CD8+ T cell responses mediate disease. Although these responses originate in the lymph node, we found that expression of the cytolytic effector molecule granzyme B was restricted to lesional CD8+ T cells in Leishmania-infected mice, suggesting that local cues within inflamed skin induced cytolytic function. Expression of Blimp-1 (Prdm1), a transcription factor necessary for cytolytic CD8+ T cell differentiation, was driven by hypoxia within the inflamed skin. Hypoxia was further enhanced by the recruitment of neutrophils that consumed oxygen to produce ROS and ultimately increased the hypoxic state and granzyme B expression in CD8+ T cells. Importantly, lesions from patients with cutaneous leishmaniasis exhibited hypoxia transcription signatures that correlated with the presence of neutrophils. Thus, targeting hypoxia-driven signals that support local differentiation of cytolytic CD8+ T cells may improve the prognosis for patients with cutaneous leishmaniasis, as well as for other inflammatory skin diseases in which cytolytic CD8+ T cells contribute to pathogenesis.
Assuntos
Linfócitos T CD8-Positivos , Leishmaniose Cutânea , Neutrófilos , Fator 1 de Ligação ao Domínio I Regulador Positivo , Animais , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/patologia , Leishmaniose Cutânea/parasitologia , Camundongos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Neutrófilos/imunologia , Neutrófilos/patologia , Neutrófilos/metabolismo , Humanos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/imunologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Granzimas/metabolismo , Granzimas/imunologia , Granzimas/genética , Hipóxia Celular/imunologia , FemininoRESUMO
Epstein-Barr virus (EBV) is deemed a necessary, yet insufficient factor in the development of multiple sclerosis (MS). In this study, myelin basic protein-specific transgenic T cell receptor mice were infected with murid gammaherpesvirus 68 virus (MHV68), an EBV-like virus that infects mice, resulting in the onset neurological deficits at a significantly higher frequency than influenza or mock-infected mice. MHV68 infected mice exhibited signs including optic neuritis and ataxia which are frequently observed in MS patients but not in experimental autoimmune encephalomyelitis mice. MHV68-infected mice exhibited increased focal immune cell infiltration in the central nervous system. Single cell RNA sequencing identified the emergence of a population of B cells that express genes associated with antigen presentation and costimulation, indicating that gammaherpesvirus infection drives a distinct, pro-inflammatory transcriptional program in B cells that may promote autoreactive T cell responses in MS.
RESUMO
Loss of cytotoxicity and defective metabolism are linked to glycogen synthase kinase 3 beta (GSK3ß) overexpression in natural killer (NK) cells from patients with acute myeloid leukemia or from healthy donors after expansion ex vivo with IL-15. Drug inhibition of GSK3ß in these NK cells improves their maturation and cytotoxic activity, but the mechanisms of GSK3ß-mediated dysfunction have not been well studied. Here, we show that expansion of NK cells with feeder cells expressing membrane-bound IL-21 maintained normal GSK3ß levels, allowing us to study GSK3ß function using CRISPR gene editing. We deleted GSK3B and expanded paired-donor knockout and wild-type (WT) NK cells and then assessed transcriptional and functional alterations induced by loss of GSK3ß. Surprisingly, our data showed that deletion of GSK3B did not alter cytotoxicity, cytokine production, or maturation (as determined by CD57 expression). However, GSK3B-KO cells demonstrated significant changes in expression of genes related to rRNA processing, cell proliferation, and metabolic function, suggesting possible metabolic reprogramming. Next, we found that key genes downregulated in GSK3B-KO NK cells were upregulated in GSK3ß-overexpressing NK cells from AML patients, confirming this correlation in a clinical setting. Lastly, we measured cellular energetics and observed that GSK3B-KO NK cells exhibited 150% higher spare respiratory capacity, a marker of metabolic fitness. These findings suggest a role for GSK3ß in regulating NK cell metabolism.