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2.
Nat Immunol ; 20(7): 824-834, 2019 07.
Article in English | MEDLINE | ID: mdl-31209403

ABSTRACT

Multiple genome-wide studies have identified associations between outcome of human immunodeficiency virus (HIV) infection and polymorphisms in and around the gene encoding the HIV co-receptor CCR5, but the functional basis for the strongest of these associations, rs1015164A/G, is unknown. We found that rs1015164 marks variation in an activating transcription factor 1 binding site that controls expression of the antisense long noncoding RNA (lncRNA) CCR5AS. Knockdown or enhancement of CCR5AS expression resulted in a corresponding change in CCR5 expression on CD4+ T cells. CCR5AS interfered with interactions between the RNA-binding protein Raly and the CCR5 3' untranslated region, protecting CCR5 messenger RNA from Raly-mediated degradation. Reduction in CCR5 expression through inhibition of CCR5AS diminished infection of CD4+ T cells with CCR5-tropic HIV in vitro. These data represent a rare determination of the functional importance of a genome-wide disease association where expression of a lncRNA affects HIV infection and disease progression.


Subject(s)
Gene Expression Regulation , Genetic Variation , HIV Infections/genetics , HIV Infections/virology , HIV-1 , RNA, Antisense/genetics , RNA, Long Noncoding/genetics , Receptors, CCR5/genetics , 3' Untranslated Regions , Alleles , Biomarkers , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD4-Positive T-Lymphocytes/virology , Cell Membrane/metabolism , Genes, Reporter , Genotype , HIV Infections/metabolism , Humans , Linkage Disequilibrium , Polymorphism, Single Nucleotide , Population Groups/genetics , Prognosis , RNA Stability , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, CCR5/metabolism , Viral Load
3.
Immunity ; 51(3): 451-464.e6, 2019 09 17.
Article in English | MEDLINE | ID: mdl-31471108

ABSTRACT

Type I and III interferons (IFNs) activate similar downstream signaling cascades, but unlike type I IFNs, type III IFNs (IFNλ) do not elicit strong inflammatory responses in vivo. Here, we examined the molecular mechanisms underlying this disparity. Type I and III IFNs displayed kinetic differences in expression of IFN-stimulated genes and proinflammatory responses, with type I IFNs preferentially stimulating expression of the transcription factor IRF1. Type III IFNs failed to induce IRF1 expression because of low IFNλ receptor abundance and insufficient STAT1 activation on epithelial cells and thus did not activate the IRF1 proinflammatory gene program. Rather, IFNλ stimulation preferentially induced factors implicated in tissue repair. Our findings suggest that IFN receptor compartmentalization and abundance confer a spatiotemporal division of labor where type III IFNs control viral spread at the site of the infection while restricting tissue damage; the transient induction of inflammatory responses by type I IFNs recruits immune effectors to promote protective immunity.


Subject(s)
Interferon Regulatory Factor-1/immunology , Interferon Type I/immunology , Interferons/immunology , Animals , Cell Line , Epithelial Cells/immunology , Humans , Inflammation/immunology , Male , Mice , Mice, Inbred C57BL , STAT1 Transcription Factor/immunology , Interferon Lambda
4.
Proc Natl Acad Sci U S A ; 119(29): e2205498119, 2022 07 19.
Article in English | MEDLINE | ID: mdl-35858344

ABSTRACT

HLA class I (HLA-I) allotypes vary widely in their dependence on tapasin (TAPBP), an integral component of the peptide-loading complex, to present peptides on the cell surface. We identified two single-nucleotide polymorphisms that regulate TAPBP messenger RNA (mRNA) expression in Africans, rs111686073 (G/C) and rs59097151 (A/G), located in an AP-2α transcription factor binding site and a microRNA (miR)-4486 binding site, respectively. rs111686073G and rs59097151A induced significantly higher TAPBP mRNA expression relative to the alternative alleles due to higher affinity for AP-2α and abrogation of miR-4486 binding, respectively. These variants associated with lower Plasmodium falciparum parasite prevalence and lower incidence of clinical malaria specifically among individuals carrying tapasin-dependent HLA-I allotypes, presumably by augmenting peptide loading, whereas tapasin-independent allotypes associated with relative protection, regardless of imputed TAPBP mRNA expression levels. Thus, an attenuated course of malaria may occur through enhanced breadth and/or magnitude of antigen presentation, an important consideration when evaluating vaccine efficacy.


Subject(s)
Histocompatibility Antigens Class I , Malaria, Falciparum , Membrane Transport Proteins , Plasmodium falciparum , Binding Sites , Genetic Variation , Histocompatibility Antigens Class I/immunology , Humans , Malaria, Falciparum/genetics , Malaria, Falciparum/immunology , Malaria, Falciparum/parasitology , Membrane Transport Proteins/genetics , Membrane Transport Proteins/metabolism , MicroRNAs/metabolism , Peptides/immunology , Plasmodium falciparum/immunology , RNA, Messenger/genetics , Transcription Factor AP-2/metabolism
5.
Immunogenetics ; 75(6): 495-506, 2023 Dec.
Article in English | MEDLINE | ID: mdl-37801092

ABSTRACT

The human KIR genes encode a family of class I MHC receptors that are expressed on subsets of NK cells. The expression of KIR proteins is controlled by a stochastic process, and competition between sense and antisense promoter elements has been suggested to program the variegated expression of these genes. Previous studies have demonstrated distinct roles of distal, intermediate, and proximal sense promoter/enhancer elements in gene activation and expression. Conversely, proximal and intronic antisense promoter transcripts have been associated with gene silencing at different stages of NK cell development. In the current study, we examine the effect of intermediate promoter deletion on KIR2DL1 expression in the YTS cell line. Homozygous deletion of the KIR2DL1 intermediate element did not affect proximal promoter activity but resulted in increased detection of upstream transcripts. No significant changes in alternative mRNA splicing or expression levels of KIR2DL1 protein were observed. However, intermediate element deletion was associated with a reduced frequency of gene activation by 5-azacytidine. Taken together, these results indicate that the intermediate element is not an enhancer required for KIR expression; however, it is required for the efficient activation of the gene.


Subject(s)
Receptors, KIR , Humans , Transcriptional Activation , Homozygote , Sequence Deletion , Receptors, KIR2DL1/genetics , Cell Line , Promoter Regions, Genetic , Receptors, KIR/genetics
6.
J Immunol ; 205(6): 1513-1523, 2020 09 15.
Article in English | MEDLINE | ID: mdl-32759296

ABSTRACT

Variegated expression of killer Ig-like receptors (KIR) in human NK cells is a stochastic process exclusive to subsets of mature NK cells and CD8+ T cells. Allele-specific KIR expression is maintained by DNA methylation within the proximal promoter regions. Because KIR genes are densely methylated in NK cell progenitors, there is an implied stage of human NK cell development in which DNA demethylation takes place to allow for active transcription. When and how this process occurs is unknown. In this study, we show that KIR proximal promoters are densely methylated in less mature CD56bright NK cells and are progressively demethylated in CD56dim NK cells as they mature and acquire KIR. We hypothesized that ten-eleven translocation (TET) enzymes, which oxidize 5mC on DNA could mediate KIR promoter demethylation. The catalytic efficiency of TET enzymes is known to be enhanced by ascorbic acid. We found that the addition of ascorbic acid to ex vivo culture of sorted CD56bright NK cells increased the frequency of KIR expression in a dose-dependent manner and facilitated demethylation of proximal promoters. A marked enrichment of the transcription factor Runx3 as well as TET2 and TET3 was observed within proximal KIR promoters in CD56bright NK cells cultured with ascorbic acid. Additionally, overexpression of TET3 and Runx3 promoted KIR expression in CD56bright NK cells and NK-92 cells. Our results show that KIR promoter demethylation can be induced in CD56bright, and this process is facilitated by ascorbic acid.


Subject(s)
Ascorbic Acid/metabolism , Killer Cells, Natural/metabolism , Receptors, KIR/metabolism , CD56 Antigen/metabolism , Cell Differentiation , Cells, Cultured , Core Binding Factor Alpha 3 Subunit/genetics , Core Binding Factor Alpha 3 Subunit/metabolism , DNA Methylation , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Demethylation , Dioxygenases/genetics , Dioxygenases/metabolism , Gene Expression Regulation , Humans , Killer Cells, Natural/immunology , Lymphocyte Activation , Promoter Regions, Genetic/genetics , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Receptors, KIR/genetics
7.
BMC Infect Dis ; 21(1): 544, 2021 Jun 09.
Article in English | MEDLINE | ID: mdl-34107889

ABSTRACT

BACKGROUND: SARS-CoV-2 is a recently emerged pandemic coronavirus (CoV) capable of causing severe respiratory illness. However, a significant number of infected people present as asymptomatic or pauci-symptomatic. In this prospective assessment of at-risk healthcare workers (HCWs) we seek to determine whether pre-existing antibody or T cell responses to previous seasonal human coronavirus (HCoV) infections affect immunological or clinical responses to SARS-CoV-2 infection or vaccination. METHODS: A cohort of 300 healthcare workers, confirmed negative for SARS-CoV-2 exposure upon study entry, will be followed for up to 1 year with monthly serology analysis of IgM and IgG antibodies against the spike proteins of SARS-CoV-2 and the four major seasonal human coronavirus - HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63. Participants will complete monthly questionnaires that ask about Coronavirus Disease 2019 (COVID-19) exposure risks, and a standardized, validated symptom questionnaire (scoring viral respiratory disease symptoms, intensity and severity) at least twice monthly and any day when any symptoms manifest. SARS-CoV-2 PCR testing will be performed any time participants develop symptoms consistent with COVID-19. For those individuals that seroconvert and/or test positive by SARS-CoV-2 PCR, or receive the SARS-CoV-2 vaccine, additional studies of T cell activation and cytokine production in response to SARS-CoV-2 peptide pools and analysis of Natural Killer cell numbers and function will be conducted on that participant's cryopreserved baseline peripheral blood mononuclear cells (PBMCs). Following the first year of this study we will further analyze those participants having tested positive for COVID-19, and/or having received an authorized/licensed SARS-CoV-2 vaccine, quarterly (year 2) and semi-annually (years 3 and 4) to investigate immune response longevity. DISCUSSION: This study will determine the frequency of asymptomatic and pauci-symptomatic SARS-CoV-2 infection in a cohort of at-risk healthcare workers. Baseline and longitudinal assays will determine the frequency and magnitude of anti-spike glycoprotein antibodies to the seasonal HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63, and may inform whether pre-existing antibodies to these human coronaviruses are associated with altered COVID-19 disease course. Finally, this study will evaluate whether pre-existing immune responses to seasonal HCoVs affect the magnitude and duration of antibody and T cell responses to SARS-CoV-2 vaccination, adjusting for demographic covariates.


Subject(s)
COVID-19/immunology , Health Personnel/statistics & numerical data , SARS-CoV-2/immunology , Seroconversion , Vaccination/statistics & numerical data , Antibodies, Viral/blood , Antibodies, Viral/immunology , Asymptomatic Infections , COVID-19 Vaccines/immunology , Coronavirus/immunology , Cross Reactions , Humans , Prospective Studies , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/immunology
8.
PLoS Genet ; 14(1): e1007163, 2018 01.
Article in English | MEDLINE | ID: mdl-29329284

ABSTRACT

The HLA-C gene appears to have evolved in higher primates to serve as a dominant source of ligands for the KIR2D family of inhibitory MHC class I receptors. The expression of NK cell-intrinsic MHC class I has been shown to regulate the murine Ly49 family of MHC class I receptors due to the interaction of these receptors with NK cell MHC in cis. However, cis interactions have not been demonstrated for the human KIR and HLA proteins. We report the discovery of an elaborate NK cell-specific system regulating HLA-C expression, indicating an important role for HLA-C in the development and function of NK cells. A large array of alternative transcripts with differences in intron/exon content are generated from an upstream NK-specific HLA-C promoter, and exon content varies between HLA-C alleles due to SNPs in splice donor/acceptor sites. Skipping of the first coding exon of HLA-C generates a subset of untranslatable mRNAs, and the proportion of untranslatable HLA-C mRNA decreases as NK cells mature, correlating with increased protein expression by mature NK cells. Polymorphism in a key Ets-binding site of the NK promoter has generated HLA-C alleles that lack significant promoter activity, resulting in reduced HLA-C expression and increased functional activity. The NK-intrinsic regulation of HLA-C thus represents a novel mechanism controlling the lytic activity of NK cells during development.


Subject(s)
HLA-C Antigens/genetics , Killer Cells, Natural/physiology , Lymphocyte Activation/genetics , Alleles , Cell Degranulation/genetics , Cells, Cultured , Gene Expression Regulation , Genes, MHC Class I , HeLa Cells , Humans , Killer Cells, Natural/immunology
9.
Int J Mol Sci ; 22(13)2021 Jun 30.
Article in English | MEDLINE | ID: mdl-34209132

ABSTRACT

The metabolic requirements and functions of cancer and normal tissues are vastly different. Due to the rapid growth of cancer cells in the tumor microenvironment, distorted vasculature is commonly observed, which creates harsh environments that require rigorous and constantly evolving cellular adaption. A common hallmark of aggressive and therapeutically resistant tumors is hypoxia and hypoxia-induced stress markers. However, recent studies have identified alterations in a wide spectrum of metabolic pathways that dictate tumor behavior and response to therapy. Accordingly, it is becoming clear that metabolic processes are not uniform throughout the tumor microenvironment. Metabolic processes differ and are cell type specific where various factors promote metabolic heterogeneity within the tumor microenvironment. Furthermore, within the tumor, these metabolically distinct cell types can organize to form cellular neighborhoods that serve to establish a pro-tumor milieu in which distant and spatially distinct cellular neighborhoods can communicate via signaling metabolites from stroma, immune and tumor cells. In this review, we will discuss how biochemical interactions of various metabolic pathways influence cancer and immune microenvironments, as well as associated mechanisms that lead to good or poor clinical outcomes.


Subject(s)
Neoplasms/immunology , Nitric Oxide/immunology , Signal Transduction/immunology , Tumor Microenvironment/immunology , Animals , Humans , Neoplasms/pathology
10.
Immunogenetics ; 72(4): 205-215, 2020 05.
Article in English | MEDLINE | ID: mdl-32219494

ABSTRACT

NK cells are primarily responsible for detecting malignant or pathogen-infected cells, and their function is influenced both by stress-associated activating signals and opposing inhibitory signals from receptors that recognize self MHC. The receptors that produce this inhibitory signal shift from the NKG2A:HLA-E system to that of KIR:HLA as the NK cells mature. This maturation is associated with an increase in lytic activity, as well as an increase in HLA-C protein levels controlled by the NK-specific HLA-C promoter, NK-Pro. We propose that modulation of the translatability of HLA-C transcripts in NK cells constitutes an evolutionary mechanism to control cis inhibitory signaling by HLA-C, which fine tunes NK cell activity. Furthermore, the high degree of variability in KIR receptor affinity for HLA alleles, as well as the variable expression levels of both KIR and HLA, suggest an evolutionary requirement for the tuning of NK lytic activity. Various data have demonstrated that mature NK cells may gain or lose lytic activity when placed in different environments. This indicates that NK cell activity may be more a function of constant tuning by inhibitory signals, rather than a static, irreversible "license to kill" granted to mature NK cells. Inhibitory signaling controls the filling of the cytolytic granule reservoir, which becomes depleted if there are insufficient inhibitory signals, leading to a hyporesponsive NK cell. We propose a novel model for the tuning of human NK cell activity via cis interactions in the context of recent findings on the mechanism of NK education.


Subject(s)
HLA-C Antigens/genetics , HLA-C Antigens/metabolism , Killer Cells, Natural/physiology , Alleles , Animals , Humans , Killer Cells, Natural/immunology , Mice , Organ Specificity , Receptors, KIR2DL1/genetics , Receptors, KIR2DL1/immunology
11.
Am J Hum Genet ; 99(6): 1353-1358, 2016 Dec 01.
Article in English | MEDLINE | ID: mdl-27817866

ABSTRACT

Differential HLA-C levels influence several human diseases, but the mechanisms responsible are incompletely characterized. Using a validated prediction algorithm, we imputed HLA-C cell surface levels in 228 individuals from the 1000 Genomes dataset. We tested 68,726 SNPs within the MHC for association with HLA-C level. The HLA-C promoter region variant, rs2395471, 800 bp upstream of the transcription start site, gave the most significant association with HLA-C levels (p = 4.2 × 10-66). This imputed expression quantitative trait locus, termed impeQTL, was also shown to associate with HLA-C expression in a genome-wide association study of 273 donors in which HLA-C mRNA expression levels were determined by quantitative PCR (qPCR) (p = 1.8 × 10-20) and in two cohorts where HLA-C cell surface levels were determined directly by flow cytometry (n = 369 combined, p < 10-15). rs2395471 is located in an Oct1 transcription factor consensus binding site motif where the A allele is predicted to have higher affinity for Oct1 than the G allele. Mobility shift electrophoresis demonstrated that Oct1 binds to both alleles in vitro, but decreased HLA-C promoter activity was observed in a luciferase reporter assay for rs2395471_G relative to rs2395471_A on a fixed promoter background. The rs2395471 variant accounts for up to 36% of the explained variation of HLA-C level. These data strengthen our understanding of HLA-C transcriptional regulation and provide a basis for understanding the potential consequences of manipulating HLA-C levels therapeutically.


Subject(s)
HLA-C Antigens/biosynthesis , HLA-C Antigens/genetics , Octamer Transcription Factor-1/metabolism , Promoter Regions, Genetic/genetics , Algorithms , Alleles , Binding Sites/genetics , Datasets as Topic , Genome, Human/genetics , Genome-Wide Association Study , HeLa Cells , Humans , Polymorphism, Single Nucleotide/genetics , Quantitative Trait Loci/genetics , Transcription Initiation Site , Transcription, Genetic
12.
13.
PLoS Biol ; 14(8): e1002526, 2016 08.
Article in English | MEDLINE | ID: mdl-27500644

ABSTRACT

It has recently been appreciated that NK cells exhibit many features reminiscent of adaptive immune cells. Considerable heterogeneity exists with respect to the ligand specificity of individual NK cells and as such, a subset of NK cells can respond, expand, and differentiate into memory-like cells in a ligand-specific manner. MHC I-binding inhibitory receptors, including those belonging to the Ly49 and KIR families, are expressed in a variegated manner, which creates ligand-specific diversity within the NK cell pool. However, how NK cells determine which inhibitory receptors to express on their cell surface during a narrow window of development is largely unknown. In this manuscript, we demonstrate that signals from activating receptors are critical for induction of Ly49 and KIR receptors during NK cell development; activating receptor-derived signals increased the probability of the Ly49 bidirectional Pro1 promoter to transcribe in the forward versus the reverse direction, leading to stable expression of Ly49 receptors in mature NK cells. Our data support a model where the balance of activating and inhibitory receptor signaling in NK cells selects for the induction of appropriate inhibitory receptors during development, which NK cells use to create a diverse pool of ligand-specific NK cells.


Subject(s)
Killer Cells, Natural/immunology , NK Cell Lectin-Like Receptor Subfamily A/immunology , Receptors, KIR/immunology , Signal Transduction/immunology , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/immunology , Adaptor Proteins, Signal Transducing/metabolism , Animals , Cells, Cultured , Flow Cytometry , Genetic Variation/immunology , Histocompatibility Antigens Class I/immunology , Histocompatibility Antigens Class I/metabolism , Humans , Killer Cells, Natural/metabolism , Ligands , Mice, Inbred C57BL , Mice, Knockout , NK Cell Lectin-Like Receptor Subfamily A/genetics , NK Cell Lectin-Like Receptor Subfamily A/metabolism , Phosphoproteins/genetics , Phosphoproteins/immunology , Phosphoproteins/metabolism , RNA Interference , Receptors, KIR/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/genetics
14.
J Immunol ; 192(8): 3889-97, 2014 Apr 15.
Article in English | MEDLINE | ID: mdl-24634493

ABSTRACT

Acquisition of a functional NK cell repertoire, known as education or licensing, is a complex process mediated through inhibitory receptors that recognize self. We found that NK cells containing self-killer Ig-like receptors for cognate HLA ligand in vivo were less susceptible to apoptosis. In vitro IL-15 withdrawal showed that uneducated NK cells upregulated Bim and Fas. Conversely, educated NK cells upregulated Fas ligand (FasL) under these conditions. Induction of cell death and Bim expression on uneducated cells correlated with increased IL-2Rα expression. Overexpression and knockdown studies showed that higher IL-2Rα limits NK cell survival in a novel manner that is independent from the role of IL-2 in activation-induced cell death. To study the role of FasL in induction of IL-2Rα(hi) NK cell death, a coculture assay with FasL-blocking Abs was used. IL-15 withdrawal led to FasL-dependent killing of IL-2Rα(hi) NK cells by more educated IL-2Rα(lo) NK cells. Finally, CMV reactivation induces a potent long-lasting population of licensed NK cells with enhanced survival. These findings show that education-dependent NK cell survival advantages and killing of uneducated NK cells result in the maintenance of a functional repertoire, which may be manipulated to exploit NK cells for cancer immunotherapy.


Subject(s)
Fas Ligand Protein/metabolism , Interleukin-2 Receptor alpha Subunit/metabolism , Killer Cells, Natural/metabolism , Animals , Apoptosis/genetics , Apoptosis/immunology , Apoptosis Regulatory Proteins/genetics , Apoptosis Regulatory Proteins/metabolism , Bcl-2-Like Protein 11 , Cell Communication , Cell Differentiation , Cell Proliferation , Cell Survival/drug effects , Cell Survival/immunology , Cytomegalovirus/physiology , Fas Ligand Protein/genetics , Gene Expression Regulation/drug effects , Homeostasis , Humans , Interleukin-15/pharmacology , Interleukin-2 Receptor alpha Subunit/genetics , Killer Cells, Natural/cytology , Killer Cells, Natural/drug effects , Killer Cells, Natural/immunology , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mice , Mice, Knockout , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Receptors, Cytokine/genetics , Receptors, Cytokine/metabolism , Receptors, KIR/genetics , Receptors, KIR/metabolism , Virus Activation
15.
J Immunol ; 192(6): 2875-84, 2014 Mar 15.
Article in English | MEDLINE | ID: mdl-24563253

ABSTRACT

Killer Ig-like receptors (KIRs) control the activation of human NK cells via interactions with peptide-laden HLAs. KIR3DL1 is a highly polymorphic inhibitory receptor that recognizes a diverse array of HLA molecules expressing the Bw4 epitope, a group with multiple polymorphisms incorporating variants within the Bw4 motif. Genetic studies suggest that KIR3DL1 variation has functional significance in several disease states, including HIV infection. However, owing to differences across KIR3DL1 allotypes, HLA-Bw4, and associated peptides, the mechanistic link with biological outcome remains unclear. In this study, we elucidated the impact of KIR3DL1 polymorphism on peptide-laden HLA recognition. Mutational analysis revealed that KIR residues involved in water-mediated contacts with the HLA-presented peptide influence peptide binding specificity. In particular, residue 282 (glutamate) in the D2 domain underpins the lack of tolerance of negatively charged C-terminal peptide residues. Allotypic KIR3DL1 variants, defined by neighboring residue 283, displayed differential sensitivities to HLA-bound peptide, including the variable HLA-B*57:01-restricted HIV-1 Gag-derived epitope TW10. Residue 283, which has undergone positive selection during the evolution of human KIRs, also played a central role in Bw4 subtype recognition by KIR3DL1. Collectively, our findings uncover a common molecular regulator that controls HLA and peptide discrimination without participating directly in peptide-laden HLA interactions. Furthermore, they provide insight into the mechanics of interaction and generate simple, easily assessed criteria for the definition of KIR3DL1 functional groupings that will be relevant in many clinical applications, including bone marrow transplantation.


Subject(s)
HLA-B Antigens/immunology , Peptides/immunology , Receptors, KIR3DL1/immunology , Amino Acid Sequence , Binding Sites/genetics , Binding Sites/immunology , Epitopes/genetics , Epitopes/immunology , HEK293 Cells , HLA-B Antigens/chemistry , HLA-B Antigens/genetics , Human Immunodeficiency Virus Proteins/genetics , Human Immunodeficiency Virus Proteins/immunology , Humans , Jurkat Cells , Killer Cells, Natural/immunology , Killer Cells, Natural/metabolism , Models, Molecular , Mutation , Peptides/chemistry , Peptides/genetics , Polymorphism, Genetic , Protein Binding/immunology , Protein Multimerization , Protein Structure, Tertiary , Receptors, KIR3DL1/chemistry , Receptors, KIR3DL1/genetics
16.
PLoS Pathog ; 9(5): e1003357, 2013 May.
Article in English | MEDLINE | ID: mdl-23675302

ABSTRACT

Fungal pathogens elicit cytokine responses downstream of immunoreceptor tyrosine-based activation motif (ITAM)-coupled or hemiITAM-containing receptors and TLRs. The Linker for Activation of B cells/Non-T cell Activating Linker (LAB/NTAL) encoded by Lat2, is a known regulator of ITAM-coupled receptors and TLR-associated cytokine responses. Here we demonstrate that LAB is involved in anti-fungal immunity. We show that Lat2-/- mice are more susceptible to C. albicans infection than wild type (WT) mice. Dendritic cells (DCs) express LAB and we show that it is basally phosphorylated by the growth factor M-CSF or following engagement of Dectin-2, but not Dectin-1. Our data revealed a unique mechanism whereby LAB controls basal and fungal/pathogen-associated molecular patterns (PAMP)-induced nuclear ß-catenin levels. This in turn is important for controlling fungal/PAMP-induced cytokine production in DCs. C. albicans- and LPS-induced IL-12 and IL-23 production was blunted in Lat2-/- DCs. Accordingly, Lat2-/- DCs directed reduced Th1 polarization in vitro and Lat2-/- mice displayed reduced Natural Killer (NK) and T cell-mediated IFN-γ production in vivo/ex vivo. Thus our data define a novel link between LAB and ß-catenin nuclear accumulation in DCs that facilitates IFN-γ responses during anti-fungal immunity. In addition, these findings are likely to be relevant to other infectious diseases that require IL-12 family cytokines and an IFN-γ response for pathogen clearance.


Subject(s)
Amino Acid Transport System y+/immunology , Candidiasis/immunology , Dendritic Cells/immunology , Fusion Regulatory Protein 1, Light Chains/immunology , Lectins, C-Type/immunology , beta Catenin/immunology , Amino Acid Transport System y+/metabolism , Animals , Candidiasis/metabolism , Dendritic Cells/metabolism , Disease Models, Animal , Female , Fusion Regulatory Protein 1, Light Chains/metabolism , Interferon-gamma/biosynthesis , Interferon-gamma/immunology , Interleukin-12/biosynthesis , Interleukin-12/immunology , Lectins, C-Type/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/immunology , beta Catenin/metabolism
17.
J Immunol ; 191(2): 688-98, 2013 Jul 15.
Article in English | MEDLINE | ID: mdl-23752612

ABSTRACT

NK subsets have activating and inhibitory receptors that bind MHC-I. Ly49A is a mouse inhibitory receptor that binds with high affinity to H2(d) in both a cis- and trans-manner. Ly49A cis-associations limit trans-interactions with H2(d)-expressing targets as well as mAb binding. We demonstrate that cis-interactions affect mAb effector functions. In vivo administration of anti-Ly49A depleted NK cells in H2(b) but not H2(d) mice. Despite lack of depletion, in vivo treatment with anti-Ly49A reduced NK killing capabilities and inhibited activation, partially due to its agonistic effect. These data explain the previously described in vivo effects on bone marrow allograft rejection observed with anti-Ly49A treatment in H2(d)-haplotype mice. However, prior treatment of mice with poly(I:C) or mouse CMV infection resulted in increased Ly49A expression and Ly49A(+) NK cell depletion in H2(d) mice. These data indicate that, although Ly49 mAbs can exert similar in vivo effects in mice with different MHC haplotypes, these effects are mediated via different mechanisms of action correlating with Ly49A expression levels and can be altered within the same strain contingent on stimuli. This illustrates the marked diversity of mAb effector functions due to the regulation of the level of expression of target Ags and responses by stimulatory incidents such as infection.


Subject(s)
Bone Marrow Cells/immunology , Bone Marrow Transplantation/immunology , Killer Cells, Natural/immunology , NK Cell Lectin-Like Receptor Subfamily A/immunology , Animals , Antibodies, Monoclonal , Cytomegalovirus/immunology , Female , Graft Rejection/immunology , Histocompatibility Antigens Class I/immunology , Histocompatibility Antigens Class I/metabolism , Killer Cells, Natural/metabolism , Mice , Mice, Inbred C57BL , Mice, Inbred DBA , NK Cell Lectin-Like Receptor Subfamily A/metabolism , Poly I-C/pharmacology
18.
Blood ; 120(17): 3455-65, 2012 Oct 25.
Article in English | MEDLINE | ID: mdl-22968455

ABSTRACT

Epistatic interactions between killer cell immunoglobulin-like receptors (KIRs) and their cognate HLA class I ligands have important implications for reproductive success, antiviral immunity, susceptibility to autoimmune conditions and cancer, as well as for graft-versus-leukemia reactions in settings of allogeneic stem cell transplantation. Although CD8 T cells are known to acquire KIRs when maturing from naive to terminally differentiated cells, little information is available about the constitution of KIR repertoires on human CD8 T cells. Here, we have performed a high-resolution analysis of KIR expression on CD8 T cells. The results show that most CD8 T cells possess a restricted KIR expression pattern, often dominated by a single activating or inhibitory KIR. Furthermore, the expression of KIR, and its modulation of CD8 T-cell function, was independent of expression of self-HLA class I ligands. Finally, despite similarities in the stochastic regulation of KIRs by the bidirectional proximal promoter, the specificity of inhibitory KIRs on CD8 T cells was often distinct from that of natural killer cells in the same individual. The results provide new insight into the formation of KIR repertoires on human T cells.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Epistasis, Genetic/immunology , Gene Expression/immunology , Killer Cells, Natural/immunology , Receptors, KIR/immunology , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/metabolism , Cell Communication/genetics , Cell Communication/immunology , Cell Differentiation/genetics , Cell Differentiation/immunology , Female , Flow Cytometry , Gene Expression Profiling , Genes, Reporter , Histocompatibility Antigens Class I/genetics , Histocompatibility Antigens Class I/immunology , Histocompatibility Antigens Class I/metabolism , Humans , Immunity, Innate , Killer Cells, Natural/cytology , Killer Cells, Natural/metabolism , Luciferases , Male , Promoter Regions, Genetic/immunology , Receptors, KIR/biosynthesis , Receptors, KIR/genetics
19.
Genes (Basel) ; 15(2)2024 02 18.
Article in English | MEDLINE | ID: mdl-38397241

ABSTRACT

An abundance of antisense promoters in the vicinity of the transcriptional start site of coding genes suggests that they play an important role in gene regulation. The divergent transcription of housekeeping genes by a common central promoter region allows for coordinated regulation of genes in related pathways and is also linked to higher promoter activity. However, closely positioned transcription start sites can also result in competition between overlapping promoter elements and generate a binary switch element. Furthermore, the direct competition resulting from the presence of an antisense promoter immediately downstream of the transcription start site of the gene produces an element that can exist in only one of two stable transcriptional states: sense or antisense. In this review, we summarize analyses of the prevalence of antisense transcription in higher eukaryotes and viruses, with a focus on the antisense promoters competing with the promoters of coding genes. The structures of bidirectional promoters driving the simultaneous expression of housekeeping genes are compared with examples of human bidirectional elements that have been shown to act as switches. Since many bidirectional elements contain a noncoding RNA as the divergent transcript, we describe examples of functional noncoding antisense transcripts that affect the epigenetic landscape and alter the expression of their host gene. Finally, we discuss opportunities for additional research on competing sense/antisense promoters, uncovering their potential role in programming cell differentiation.


Subject(s)
Genome, Human , Transcription, Genetic , Humans , Prevalence , Promoter Regions, Genetic , Gene Expression Regulation/genetics
20.
JCI Insight ; 9(12)2024 May 21.
Article in English | MEDLINE | ID: mdl-38912586

ABSTRACT

Immune therapy is the new frontier of cancer treatment. Therapeutic radiation is a known inducer of immune response and can be limited by immunosuppressive mediators including cyclooxygenase-2 (COX2) that is highly expressed in aggressive triple negative breast cancer (TNBC). A clinical cohort of TNBC tumors revealed poor radiation therapeutic efficacy in tumors expressing high COX2. Herein, we show that radiation combined with adjuvant NSAID (indomethacin) treatment provides a powerful combination to reduce both primary tumor growth and lung metastasis in aggressive 4T1 TNBC tumors, which occurs in part through increased antitumor immune response. Spatial immunological changes including augmented lymphoid infiltration into the tumor epithelium and locally increased cGAS/STING1 and type I IFN gene expression were observed in radiation-indomethacin-treated 4T1 tumors. Thus, radiation and adjuvant NSAID treatment shifts "immune desert phenotypes" toward antitumor M1/TH1 immune mediators in these immunologically challenging tumors. Importantly, radiation-indomethacin combination treatment improved local control of the primary lesion, reduced metastatic burden, and increased median survival when compared with radiation treatment alone. These results show that clinically available NSAIDs can improve radiation therapeutic efficacy through increased antitumor immune response and augmented local generation of cGAS/STING1 and type I IFNs.


Subject(s)
Membrane Proteins , Signal Transduction , T-Lymphocytes, Cytotoxic , Animals , Membrane Proteins/metabolism , Mice , Female , Signal Transduction/drug effects , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/drug effects , Triple Negative Breast Neoplasms/pathology , Triple Negative Breast Neoplasms/immunology , Triple Negative Breast Neoplasms/drug therapy , Triple Negative Breast Neoplasms/radiotherapy , Indomethacin/pharmacology , Indomethacin/therapeutic use , Cell Line, Tumor , Humans , Lung Neoplasms/immunology , Lung Neoplasms/pathology , Lung Neoplasms/drug therapy , Cyclooxygenase Inhibitors/pharmacology , Cyclooxygenase Inhibitors/therapeutic use , Nucleotidyltransferases/metabolism , Interferon Type I/metabolism , Cyclooxygenase 2/metabolism , Lymphocytes, Tumor-Infiltrating/immunology , Lymphocytes, Tumor-Infiltrating/drug effects , Mice, Inbred BALB C
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