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1.
Scand J Immunol ; 90(4): e12797, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31166602

RESUMO

Distinctive "two signal" paths in immunology, taken by researchers with different academic backgrounds, seem to have both contained facets of the truth. Having been influenced by education at a medical school where Almroth Wright's early contributions were not forgotten, the author's "path less followed" led to views that began to gain recognition late in the twentieth century when the intimate relationship between innate and acquired immunity became more apparent.


Assuntos
Modelos Imunológicos , Agregação de Receptores , Linfócitos T/fisiologia , Timo/imunologia , Imunidade Adaptativa , Animais , Autoantígenos/imunologia , Autoimunidade , Consenso , Quadruplex G , Humanos , Imunidade Inata , Ativação Linfocitária , Tolerância a Antígenos Próprios
2.
Immunobiology ; 224(3): 362-370, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30876792

RESUMO

Given the heightened interest in manipulation of co-signaling cascades for cancer immunotherapy, we sought to determine how/whether tumors decorated with therapeutic monoclonal antibodies (mAbs) impact the expression of co-signaling molecules on human NK cells. Stimulation of NK cells with aggregated IgG1 resulted in the upregulation of HAVCR2 - the gene encoding T-cell immunoglobulin and mucin-containing domain (Tim)-3 - known to be involved in the induction of peripheral T cell tolerance. This upregulation of HAVCR2 was recapitulated at the protein level, following NK cell stimulation by either mAb opsonized tumors, recombinant human IgG1 Fc multimer, and/or non-Fc stimuli e.g. IL-12/IL-18. The patterns of Tim-3 expression were temporally distinct from the FcR mediated induction of the co-signaling molecule, 4-1BB (CD137), with Tim-3 increases observed twenty minutes following exposure to Fc multimers and remaining at high levels for at least six hours, while increases in CD137 expression were first observed at the four-hour time point. Importantly, these Tim-3+ NK cells were functionally diverse, as evidenced by the fact that their ability to produce IFN-γ in response to an NK cell responsive tumor was strictly dependent upon the stimuli employed for Tim-3 induction. These data suggest that Tim-3 upregulation is the common end-result of NK cell activation by a variety of unique and overlapping stimuli and is not an independent marker of NK cell exhaustion. Furthermore, our observations potentially explain the diverse functionality attributed to Tim-3+ NK cells and should be considered prior to use of anti-Tim-3 inhibitory mAbs for cancer immunotherapy.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Imunoglobulina G/metabolismo , Imunoterapia/métodos , Células Matadoras Naturais/imunologia , Neoplasias/terapia , Células Cultivadas , Receptor Celular 2 do Vírus da Hepatite A/genética , Receptor Celular 2 do Vírus da Hepatite A/imunologia , Humanos , Tolerância Imunológica , Interferon gama/metabolismo , Células K562 , Ativação Linfocitária , Neoplasias/imunologia , Multimerização Proteica , Agregação de Receptores , Receptores Fc/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Regulação para Cima
3.
Front Immunol ; 9: 2085, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30279692

RESUMO

We created APC-mimetic synthetic substrates to study the impact of ligand clustering on T cell activation and spreading. The substrates exhibit antibodies directed against the TCR-complex in the form of a patterned array of sub micrometric dots surrounded by a fluid supported lipid bilayer (SLB) which may itself be functionalized with another bio-molecule. We show that for T cell adhesion mediated by T cell receptor (TCR) alone, in the patterned, but not in the corresponding homogeneous controls, the TCR, ZAP-70 and actin are present in the form of clusters or patches that co-localize with the ligand-dots. However, global cell scale parameters like cell area and actin distribution are only weakly impacted by ligand clustering. In presence of ICAM-1 - the ligand of the T cell integrin LFA-1 - on the SLB, the TCR is still clustered due to the patterning of its ligands, but now global parameters are also impacted. The actin organization changes to a peripheral ring, resembling the classical actin distribution seen on homogeneous substrates, the patterned membrane topography disappears and the membrane is flat, whereas the cell area increases significantly. These observations taken together point to a possible pivotal role for LFA-1 in amplifying the effect of TCR-clustering. No such effect is evident for co-engagement of CD28, affected via its ligand B7.2. Unlike on ICAM-1, on B7.2 cell spreading and actin organization are similar for homogeneous and patterned substrates. However, TCR and ZAP-70 clusters are still formed in the patterned case. These results indicate complementary role for LFA-1 and CD28 in the regulation and putative coupling of TCR micro-clusters to actin. The engineered substrates presented here clearly have the potential to act as platform for fundamental research in immune cell biology, as well as translational analyses in immunotherapy, for example to screen molecules for their role in T cell adhesion/activation.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Biomimética/métodos , Antígeno-1 Associado à Função Linfocitária/metabolismo , Complexos Multiproteicos/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/imunologia , Actinas/metabolismo , Células Apresentadoras de Antígenos/química , Antígenos CD28/metabolismo , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Células Jurkat , Bicamadas Lipídicas/química , Ativação Linfocitária , Agregação de Receptores , Receptor Cross-Talk , Transdução de Sinais , Proteína-Tirosina Quinase ZAP-70/metabolismo
4.
Immunol Res ; 66(5): 557-566, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30269202

RESUMO

Leptin, the adipose tissue-derived product of the obese (ob) gene, is known to function as the hormone of energy expenditure. It has also been established that leptin regulates immune and inflammatory processes. All leptin-induced biological activities depend on binding to the membrane-spanning leptin receptor (Ob-R), belonging to the class I cytokine receptor family. The available data relating to the Ob-R on mature mast cells (MCs), and consequently leptin significance in the modulation of MC activity within the tissue, are limited. Immunohistochemistry was used to establish Ob-R expression by MCs in the mesenteric adipose tissue. Flow cytometry and confocal microscopy were used to evaluate both constitutive and leptin-induced expression of Ob-R on freshly isolated peritoneal MCs. MCs in the mesenteric adipose tissue and native peritoneal MCs express Ob-R constitutively. Additionally, leptin influences its receptor expression on these cells. Leptin at lower concentrations caused Ob-R expression increase both at the cell surface and in the cell interior. MC stimulation with higher concentrations of leptin results in a decline of Ob-R from the cell surface and significant enhancement of this receptor not only in the nuclear region but also in the endoplasmic reticulum. In conclusion, one can be assumed that leptin regulates MC activity within tissues. These findings might provide an additional link among the leptin, innate immune function, and inflammatory processes and diseases.


Assuntos
Tecido Adiposo/citologia , Mastócitos/imunologia , Receptores para Leptina/metabolismo , Animais , Células Cultivadas , Metabolismo Energético , Feminino , Imunidade Inata , Leptina/metabolismo , Mesentério/citologia , Peritônio/citologia , Transporte Proteico , Ratos , Ratos Wistar , Agregação de Receptores
5.
Nat Immunol ; 19(9): 1001-1012, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30104633

RESUMO

Immunoglobulin G3 (IgG3) has an uncertain role in the response to infection with and vaccination against human immunodeficiency virus (HIV). Here we describe a regulatory role for IgG3 in dampening the immune system-activating effects of chronic HIV viremia on B cells. Secreted IgG3 was bound to IgM-expressing B cells in vivo in HIV-infected chronically viremic individuals but not in early-viremic or aviremic individuals. Tissue-like memory (TLM) B cells, a population expanded by persistent HIV viremia, bound large amounts of IgG3. IgG3 induced clustering of B cell antigen receptors (BCRs) on the IgM+ B cells, which was mediated by direct interactions between soluble IgG3 and membrane IgM of the BCR (IgM-BCR). The inhibitory IgG receptor CD32b (FcγRIIb), complement component C1q and inflammatory biomarker CRP contributed to the binding of secreted IgG3 onto IgM-expressing B cells of HIV-infected individuals. Notably, IgG3-bound TLM B cells were refractory to IgM-BCR stimulation, thus demonstrating that IgG3 can regulate B cells during chronic activation of the immune system.


Assuntos
Linfócitos B/imunologia , Infecções por HIV/imunologia , HIV-1/fisiologia , Imunoglobulina G/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Adulto , Proteína C-Reativa/metabolismo , Células Cultivadas , Complemento C1q/metabolismo , Feminino , Humanos , Imunoglobulina M/metabolismo , Memória Imunológica , Imunomodulação , Masculino , Pessoa de Meia-Idade , Ligação Proteica , Agregação de Receptores , Receptores de IgG/metabolismo , Adulto Jovem
6.
Nat Immunol ; 19(8): 821-827, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-30013143

RESUMO

The main function of T cells is to identify harmful antigens as quickly and precisely as possible. Super-resolution microscopy data have indicated that global clustering of T cell antigen receptors (TCRs) occurs before T cell activation. Such pre-activation clustering has been interpreted as representing a potential regulatory mechanism that fine tunes the T cell response. We found here that apparent TCR nanoclustering could be attributed to overcounting artifacts inherent to single-molecule-localization microscopy. Using complementary super-resolution approaches and statistical image analysis, we found no indication of global nanoclustering of TCRs on antigen-experienced CD4+ T cells under non-activating conditions. We also used extensive simulations of super-resolution images to provide quantitative limits for the degree of randomness of the TCR distribution. Together our results suggest that the distribution of TCRs on the plasma membrane is optimized for fast recognition of antigen in the first phase of T cell activation.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Membrana Celular/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Animais , Células Cultivadas , Senescência Celular , Simulação por Computador , Memória Imunológica , Ativação Linfocitária , Camundongos , Camundongos Transgênicos , Imagens de Fantasmas , Ligação Proteica , Agregação de Receptores , Receptores de Antígenos de Linfócitos T alfa-beta/genética
7.
Front Immunol ; 9: 415, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29552015

RESUMO

CCR6 is a G protein-coupled receptor (GPCR) that recognizes a single chemokine ligand, CCL20 and is primarily expressed by leukocytes. Upon ligand binding, CCR6 activates Gαi heterotrimeric G proteins to induce various potential cellular outcomes through context-specific cell signaling. It is well known that differential phosphorylation of Ser and Thr residues in the C-terminal domains or intracellular loops of GPCRs can generate barcodes that regulate GPCR function by regulating the recruitment of ß-arrestins. In this study, we demonstrate that ligand binding to CCR6 induces receptor phosphorylation at Ser/Thr residues in the C-terminal tail, rather than intracellular loops. Using mutagenesis experiments, we determined that distinct clusters of Ser/Thr residues in the C-terminal domain differentially regulate CCL20-induced signaling and cellular response. Substituting the Thr360/Ser361/Thr363 cluster or the Ser370/Ser371 cluster with Ala residues modulated cellular response upon CCL20 stimulation. Notably, receptor internalization, chemotaxis, F-actin distribution, transient ERK1/2 activation, and ß-arrestin 2 recruitment were oppositely affected by mutating the two clusters, suggesting that phosphorylation of CCR6 C-terminal Ser/Thr residues directs the cell signaling response upon receptor activation. Moreover, activated CCR6 weakly recruited ß-arrestin 1 in comparison with ß-arrestin 2, and the two arrestin proteins seemed to play overlapping but distinct roles in mediating CCL20/CCR6-induced cellular responses. Taken together, the effects of site-specific Ser/Thr phosphorylation on CCR6 demonstrate the existence of barcodes on the protein that dictate the activation of different cell signaling profiles and lead to distinct biological outcomes.


Assuntos
Actinas/metabolismo , Receptores CCR6/metabolismo , Transdução de Sinais/genética , Humanos , Células Jurkat , Mutagênese Sítio-Dirigida , Mutação/genética , Fosforilação , Domínios Proteicos/genética , Agregação de Receptores/genética , Receptores CCR6/genética , Serina/genética , Serina/metabolismo , Treonina/genética , Treonina/metabolismo , beta-Arrestina 1/metabolismo , beta-Arrestina 2/metabolismo
8.
Immunity ; 47(1): 159-170.e10, 2017 07 18.
Artigo em Inglês | MEDLINE | ID: mdl-28723548

RESUMO

Clearance of pathogens or tumor cells by antibodies traditionally requires both Fab and Fc domains of IgG. Here, we show the Fc domain of IgG alone mediates recognition and clearance of herpes simplex virus (HSV1)-infected cells. The human natural killer (NK) cell surface is naturally coated with IgG bound by its Fc domain to the Fcγ receptor CD16a. NK cells utilize the Fc domain of bound IgG to recognize gE, an HSV1-encoded glycoprotein that also binds the Fc domain of IgG but at a site distinct from CD16a. The bridge formed by the Fc domain between the HSV1-infected cell and the NK cell results in NK cell activation and lysis of the HSV1-infected cell in the absence of HSV1-specific antibody in vitro and prevents fatal HSV1 infection in vivo. This mechanism also explains how bacterial IgG-binding proteins regulate NK cell function and may be broadly applicable to Fcγ-receptor-bearing cells.


Assuntos
Anticorpos Antivirais/metabolismo , Herpes Simples/imunologia , Fragmentos Fc das Imunoglobulinas/metabolismo , Imunoglobulina G/metabolismo , Células Matadoras Naturais/imunologia , Simplexvirus/imunologia , Animais , Anticorpos Antivirais/imunologia , Células Cultivadas , Citotoxicidade Imunológica , Feminino , Humanos , Fragmentos Fc das Imunoglobulinas/imunologia , Imunoglobulina G/imunologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Ligação Proteica , Agregação de Receptores , Receptores de IgG/metabolismo , Transdução de Sinais , Proteínas Virais/imunologia
9.
J Immunol ; 199(5): 1817-1826, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28739877

RESUMO

PALLD is an actin cross-linker supporting cellular mechanical tension. However, its involvement in the regulation of phagocytosis, a cellular activity essential for innate immunity and physiological tissue turnover, is unclear. We report that PALLD is highly induced along with all-trans-retinoic acid-induced maturation of myeloid leukemia cells, to promote Ig- or complement-opsonized phagocytosis. PALLD mechanistically facilitates phagocytic receptor clustering by regulating actin polymerization and c-Src dynamic activation during particle binding and early phagosome formation. PALLD is also required at the nascent phagosome to recruit phosphatase oculocerebrorenal syndrome of Lowe, which regulates phosphatidylinositol-4,5-bisphosphate hydrolysis and actin depolymerization to complete phagosome closure. Collectively, our results show a new function for PALLD as a crucial regulator of the early phase of phagocytosis by elaborating dynamic actin polymerization and depolymerization.


Assuntos
Actinas/metabolismo , Proteínas do Citoesqueleto/metabolismo , Células Dendríticas/imunologia , Leucemia Mieloide Aguda/imunologia , Células-Tronco Neoplásicas/fisiologia , Síndrome Oculocerebrorrenal/imunologia , Fagocitose , Fosfoproteínas/metabolismo , Animais , Diferenciação Celular , Linhagem Celular Tumoral , Autorrenovação Celular , Proteínas do Citoesqueleto/genética , Humanos , Imunidade Inata , Camundongos , Camundongos Endogâmicos C57BL , Fagossomos/metabolismo , Fosfoproteínas/genética , Monoéster Fosfórico Hidrolases/metabolismo , Polimerização , Agregação de Receptores , Tretinoína/metabolismo
10.
J Math Biol ; 75(3): 705-731, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28124076

RESUMO

In this paper we construct and analyze a model of cell receptor aggregation. Experiments have shown that receptors in an aggregated state have greatly reduced mobility. We model the effects of this reduced mobility with a density dependent diffusion and study the impact of density dependent diffusion on aggregate formation in a one-dimensional domain. Critical values of receptor diffusivity and receptor activation are found and compared with numerical simulations. We find that the role of density dependant diffusion is quite limited in the formation of aggregate structures. In the case of receptor activation, the analytical results agree very well with the numerical calculations. Finally, we consider our model in higher dimensional domains. In this case our analysis is primarily numerical.


Assuntos
Modelos Biológicos , Agregação de Receptores , Receptores de Superfície Celular , Difusão
11.
Nat Immunol ; 18(2): 214-224, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27992402

RESUMO

The signaling adaptor MAVS forms prion-like aggregates to activate an innate antiviral immune response after viral infection. However, the molecular mechanisms that regulate MAVS aggregation are poorly understood. Here we identified TRIM31, an E3 ubiquitin ligase of the TRIM family of proteins, as a regulator of MAVS aggregation. TRIM31 was recruited to mitochondria after viral infection and specifically regulated antiviral signaling mediated by RLR pattern-recognition receptors. TRIM31-deficient mice were more susceptible to infection with RNA virus than were wild-type mice. TRIM31 interacted with MAVS and catalyzed the Lys63 (K63)-linked polyubiquitination of Lys10, Lys311 and Lys461 on MAVS. This modification promoted the formation of prion-like aggregates of MAVS after viral infection. Our findings reveal new insights in the molecular regulation of MAVS aggregation and the cellular antiviral response through TRIM31-mediated K63-linked polyubiquitination of MAVS.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Transporte/metabolismo , Macrófagos/fisiologia , Proteínas Nucleares/metabolismo , Príons/imunologia , Viroses/imunologia , Animais , Proteínas de Transporte/genética , Células Cultivadas , Imunidade Inata/genética , Lisina/genética , Lisina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Nucleares/genética , Agregação de Receptores/genética , Transdução de Sinais/genética , Ubiquitinação/genética
12.
Transpl Immunol ; 40: 22-30, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28017877

RESUMO

Donor-specific antibody (DSA), particularly against HLA class II, is a major cause of chronic antibody-mediated rejection (CAMR) after transplantation, although ABO-incompatible kidney transplantation has recently demonstrated favorable graft outcomes. The condition of no injury even in the presence of anti-donor antibody has been referred to as "accommodation", which would be one of the key factors for successful long-term graft survival. The purpose of this study was to analyze the beneficial effect of anti-blood group A/B antibody ligation on endothelial cells against HLA-DR antibody-mediated, complement-dependent cytotoxicity (CDC). Blood group A/B-expressing endothelial cells EA.hy926 or Human Umbilical Vein Endothelia Cells (HUVEC) were incubated with IFNγ in the presence or absence of anti-blood group A/B antibody or mTOR inhibitor (mTOR-i) for 48h. The effects on signaling pathway, HLA expression, complement regulatory factors, and CDC were investigated. Expression of HLA-DR on EA.hy926 or HUVEC were successfully elicited by IFNγ treatment, although little or no expression was observed in quiescent cells. Pre-incubation with anti-blood group A/B antibody had resistance to HLA-DR antibody-mediated CDC against IFNγ-treated cells in a concentration-dependent manner. This finding was ascribed to decreased expression of HLA-DR by post-translational regulation and increased expression of CD55/59, which was related to ERK and mTOR pathway inhibition. mTOR-i also inhibited HLA-DR expression by itself. Furthermore, the combination of mTOR-I and anti-blood group A/B ligation had an additive effect in preventing HLA-DR antibody-mediated CDC. Anti-blood group A/B antibody might play a preventive role in CAMR. Inhibition of the ERK and mTOR pathways may contribute to the development of a novel treatment in the maintenance period after transplantation.


Assuntos
Sistema ABO de Grupos Sanguíneos/imunologia , Anticorpos/farmacologia , Células Endoteliais/imunologia , Everolimo/farmacologia , Antígenos HLA-DR/metabolismo , Transplante de Rim , Serina-Treonina Quinases TOR/antagonistas & inibidores , Citotoxicidade Celular Dependente de Anticorpos , Células Endoteliais/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação da Expressão Gênica , Antígenos HLA-DR/genética , Antígenos HLA-DR/imunologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Interferon gama/imunologia , Processamento de Proteína Pós-Traducional , Agregação de Receptores , Transdução de Sinais , Tolerância ao Transplante , Transplante Homólogo
13.
Trends Immunol ; 37(11): 790-802, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27667711

RESUMO

NKG2D is an activating receptor that can bind to a large number of stress-induced ligands that are expressed in the context of cancer or viral infection. This receptor is expressed on many cytotoxic lymphocytes, and plays a crucial role in antitumor and antiviral immune responses. However, exposure to NKG2D ligand-expressing target cells promotes receptor endocytosis, ultimately leading to lysosomal receptor degradation and impairment of NKG2D-mediated functions. Interestingly, before being degraded, internalized receptors can signal from the endosomal compartment, leading to the appropriate activation of cellular functional programs. This review summarizes recent findings on ligand-induced receptor internalization, with particular emphasis on the role of endocytosis in the control of both NKG2D-mediated intracellular signaling and receptor degradation.


Assuntos
Endocitose/imunologia , Células Matadoras Naturais/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Neoplasias/imunologia , Viroses/imunologia , Animais , Regulação da Expressão Gênica , Humanos , Ativação Linfocitária , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Agregação de Receptores , Transdução de Sinais/imunologia , Estresse Fisiológico
14.
Curr Opin Immunol ; 42: 113-118, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27619413

RESUMO

For proteins to become allergenic, they need to acquire features enabling them to induce B cell activation and isotype switch to IgE production. Crosslinking of the B-cell receptor (BCR) is the most efficient way to productively activate B-cells. The IgE-crosslinking capability of allergens is equally crucial in the effector phase of immediate type allergy. Antigens, which acquire enhanced crosslinking capacity by oligomerization, aggregation, or the expression of repetitive epitopes may therefore gain allergenic potency. The accumulated evidence for repetitive epitope display by allergens suggests the existence of allergen-associated molecular patterns.


Assuntos
Alérgenos/imunologia , Linfócitos B/imunologia , Epitopos de Linfócito B/imunologia , Hipersensibilidade Imediata/imunologia , Padrões Moleculares Associados a Patógenos/imunologia , Proteínas de Plantas/imunologia , Receptores de Antígenos de Linfócitos B/metabolismo , Animais , Humanos , Imunoglobulina E/biossíntese , Ativação Linfocitária , Agregação de Receptores
15.
Oncotarget ; 7(40): 64942-64956, 2016 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-27409341

RESUMO

DR4 (Death Receptor 4) and DR5 (Death Receptor 5) are two potential targets for cancer therapy due to their ability to trigger apoptosis of cancer cells, but not normal ones, when activated by their cognate ligand TRAIL (TNF related apoptosis-inducing ligand). Therapies based on soluble recombinant TRAIL or agonist antibodies directed against one of the receptors are currently under clinical trials. However, TRAIL-R positive tumor cells are frequently resistant to TRAIL induced apoptosis. The precise mechanisms of this resistance are still not entirely understood. We have previously reported on synthetic peptides that bind to DR5 (TRAILmim/DR5) and induce tumor cell apoptosis in vitro and in vivo. Here, we showed that while hexameric soluble TRAIL is able to efficiently kill the DR5 positive lymphoma Jurkat or the carcinoma HCT116, these cells are resistant to apoptosis induced by the divalent form of TRAILmim/DR5 and are poorly sensitive to apoptosis induced by an anti-DR5 agonist monoclonal antibody. This resistance can be restored by the cross-linking of anti-DR5 agonist antibody but not by the cross-linking of the divalent form of TRAILmim/DR5. Interestingly, the divalent form of TRAILmim/DR5 that induced apoptosis of DR5 positive BJAB cells, acts as an inhibitor of TRAIL-induced apoptosis on Jurkat and HCT116 cells. The rapid internalization of DR5 observed when treated with divalent form of TRAILmim/DR5 could explain the antagonist activity of the ligand on Jurkat and HCT116 cells but also highlights the independence of the mechanisms responsible for internalization and activation when triggering the DR5 apoptotic cascade.


Assuntos
Imunoterapia/métodos , Neoplasias/metabolismo , Multimerização Proteica , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Anticorpos Monoclonais/química , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais Humanizados , Apoptose , Células HCT116 , Humanos , Células Jurkat , Ligantes , Terapia de Alvo Molecular , Neoplasias/terapia , Especificidade de Órgãos , Agregação de Receptores , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/imunologia , Ligante Indutor de Apoptose Relacionado a TNF/síntese química , Ligante Indutor de Apoptose Relacionado a TNF/uso terapêutico
16.
EBioMedicine ; 9: 207-216, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27333049

RESUMO

Plasmodium falciparum malaria is a deadly pathogen. The invasion of red blood cells (RBCs) by merozoites is a target for vaccine development. Although anti-merozoite antibodies can block invasion in vitro, there is no efficacy in vivo. To explain this discrepancy we hypothesized that complement activation could enhance RBC invasion by binding to the complement receptor 1 (CR1). Here we show that a monoclonal antibody directed against the merozoite and human polyclonal IgG from merozoite vaccine recipients enhanced RBC invasion in a complement-dependent manner and that soluble CR1 inhibited this enhancement. Sialic acid-independent strains, that presumably are able to bind to CR1 via a native ligand, showed less complement-dependent enhancement of RBC invasion than sialic acid-dependent strains that do not utilize native CR1 ligands. Confocal fluorescent microscopy revealed that complement-dependent invasion resulted in aggregation of CR1 at the RBC surface in contact with the merozoite. Finally, total anti-P. berghei IgG enhanced parasite growth and C3 deficiency decreased parasite growth in mice. These results demonstrate, contrary to current views, that complement activation in conjunction with antibodies can paradoxically aid parasites invade RBCs and should be considered in future design and testing of merozoite vaccines.


Assuntos
Anticorpos Monoclonais/imunologia , Imunoglobulina G/imunologia , Plasmodium falciparum/patogenicidade , Receptores de Complemento/metabolismo , Animais , Anticorpos Monoclonais/farmacologia , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Eritrócitos/citologia , Eritrócitos/metabolismo , Eritrócitos/parasitologia , Citometria de Fluxo , Humanos , Malária/parasitologia , Merozoítos/efeitos dos fármacos , Merozoítos/imunologia , Merozoítos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Confocal , Parasitemia/parasitologia , Parasitemia/patologia , Parasitemia/veterinária , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/metabolismo , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/metabolismo , Vacinas Protozoárias/imunologia , Agregação de Receptores , Receptores de Complemento/química
17.
J Immunol ; 196(9): 3951-62, 2016 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-27036914

RESUMO

Sphingolipids are major components of the plasma membrane. In particular, ceramide serves as an essential building hub for complex sphingolipids, but also as an organizer of membrane domains segregating receptors and signalosomes. Sphingomyelin breakdown as a result of sphingomyelinase activation after ligation of a variety of receptors is the predominant source of ceramides released at the plasma membrane. This especially applies to T lymphocytes where formation of ceramide-enriched membrane microdomains modulates TCR signaling. Because ceramide release and redistribution occur very rapidly in response to receptor ligation, novel tools to further study these processes in living T cells are urgently needed. To meet this demand, we synthesized nontoxic, azido-functionalized ceramides allowing for bio-orthogonal click-reactions to fluorescently label incorporated ceramides, and thus investigate formation of ceramide-enriched domains. Azido-functionalized C6-ceramides were incorporated into and localized within plasma membrane microdomains and proximal vesicles in T cells. They segregated into clusters after TCR, and especially CD28 ligation, indicating efficient sorting into plasma membrane domains associated with T cell activation; this was abolished upon sphingomyelinase inhibition. Importantly, T cell activation was not abrogated upon incorporation of the compound, which was efficiently excluded from the immune synapse center as has previously been seen in Ab-based studies using fixed cells. Therefore, the functionalized ceramides are novel, highly potent tools to study the subcellular redistribution of ceramides in the course of T cell activation. Moreover, they will certainly also be generally applicable to studies addressing rapid stimulation-mediated ceramide release in living cells.


Assuntos
Azidas/metabolismo , Microdomínios da Membrana/metabolismo , Microscopia de Fluorescência/métodos , Receptores de Antígenos de Linfócitos T/metabolismo , Esfingolipídeos/metabolismo , Linfócitos T/metabolismo , Azidas/química , Células Cultivadas , Humanos , Ativação Linfocitária , Transporte Proteico , Agregação de Receptores , Transdução de Sinais , Esfingolipídeos/química , Linfócitos T/imunologia
18.
J Immunol ; 196(6): 2767-78, 2016 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-26864032

RESUMO

ICAM-1 is required for firm adhesion of leukocytes to the endothelium. However, how the spatial organization of endothelial ICAM-1 regulates leukocyte adhesion is not well understood. In this study, we identified the calcium-effector protein annexin A2 as a novel binding partner for ICAM-1. ICAM-1 clustering promotes the ICAM-1-annexin A2 interaction and induces translocation of ICAM-1 into caveolin-1-rich membrane domains. Depletion of endothelial annexin A2 using RNA interference enhances ICAM-1 membrane mobility and prevents the translocation of ICAM-1 into caveolin-1-rich membrane domains. Surprisingly, this results in increased neutrophil adhesion and transendothelial migration under flow conditions and reduced crawling time, velocity, and lateral migration distance of neutrophils on the endothelium. In conclusion, our data show that annexin A2 limits neutrophil transendothelial migration by organizing the spatial distribution of ICAM-1.


Assuntos
Anexina A2/metabolismo , Células Endoteliais/imunologia , Molécula 1 de Adesão Intercelular/metabolismo , Leucócitos/imunologia , Neutrófilos/imunologia , Caveolina 1/metabolismo , Adesão Celular , Movimento Celular , Células Endoteliais da Veia Umbilical Humana , Humanos , Ligação Proteica , Transporte Proteico , Agregação de Receptores , Migração Transendotelial e Transepitelial
19.
Annu Rev Immunol ; 34: 243-64, 2016 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-26907217

RESUMO

Galectins are a family of mammalian carbohydrate-binding proteins expressed by many cell types. Galectins can function intracellularly and can also be secreted to bind to cell surface glycoconjugate counterreceptors. Some galectins are made by immune cells, whereas other galectins are secreted by different cell types, such as endothelial or epithelial cells, and bind to immune cells to regulate immune responses. Galectin binding to a single glycan ligand is a low-affinity interaction, but the multivalency of galectins and the glycan ligands presented on cell surface glycoproteins results in high-avidity binding that can reversibly scaffold or cluster these glycoproteins. Galectin binding to a specific glycoprotein counterreceptor is regulated in part by the repertoire of glycosyltransferase enzymes (which make the glycan ligands) expressed by that cell, and the effect of galectin binding results from clustering or retention of specific glycoprotein counterreceptors bearing these specific ligands.


Assuntos
Galectinas/metabolismo , Glicosiltransferases/metabolismo , Imunidade , Animais , Carboidratos/imunologia , Citoesqueleto , Galectinas/imunologia , Glicoproteínas/metabolismo , Humanos , Ligação Proteica , Agregação de Receptores
20.
J Biol Chem ; 291(7): 3174-83, 2016 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-26719327

RESUMO

Several different receptor proteins have been identified that bind monomeric, oligomeric, or fibrillar forms of amyloid-ß (Aß). "Good" receptors internalize Aß or promote its transcytosis out of the brain, whereas "bad" receptors bind oligomeric forms of Aß that are largely responsible for the synapticloss, memory impairments, and neurotoxicity that underlie Alzheimer disease. The prion protein both removes Aß from the brain and transduces the toxic actions of Aß. The clustering of distinct receptors in cell surface signaling platforms likely underlies the actions of distinct oligomeric species of Aß. These Aß receptor-signaling platforms provide opportunities for therapeutic intervention in Alzheimer disease.


Assuntos
Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Modelos Biológicos , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Receptores de Superfície Celular/agonistas , Transdução de Sinais , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/patologia , Doença de Alzheimer/prevenção & controle , Peptídeos beta-Amiloides/antagonistas & inibidores , Peptídeos beta-Amiloides/química , Animais , Apoptose/efeitos dos fármacos , Humanos , Ligantes , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/agonistas , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Terapia de Alvo Molecular , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/química , Neurônios/efeitos dos fármacos , Neurônios/patologia , Nootrópicos/farmacologia , Nootrópicos/uso terapêutico , Proteínas PrPC/agonistas , Proteínas PrPC/antagonistas & inibidores , Proteínas PrPC/metabolismo , Agregados Proteicos/efeitos dos fármacos , Agregação Patológica de Proteínas/tratamento farmacológico , Agregação Patológica de Proteínas/metabolismo , Agregação Patológica de Proteínas/patologia , Agregação Patológica de Proteínas/prevenção & controle , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Agregação de Receptores/efeitos dos fármacos , Receptor para Produtos Finais de Glicação Avançada/agonistas , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Receptores de Superfície Celular/antagonistas & inibidores , Receptores de Superfície Celular/metabolismo , Receptores de N-Metil-D-Aspartato/agonistas , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transcitose/efeitos dos fármacos
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