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1.
Blood ; 2021 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-33512478

RESUMO

Dendritic cells (DCs) encompass several cell subsets that collaborate to initiate and regulate immune responses. Proper DC localization determines their function and requires the tightly controlled action of chemokine receptors. All DC subsets express CXCR4, but the genuine contribution of this receptor to their biology has been overlooked. We addressed this question using natural CXCR4 mutants resistant to CXCL12-induced desensitization and harboring a gain of function that cause the warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) Syndrome (WS), a rare immunodeficiency associated with high susceptibility to the pathogenesis of human papillomavirus (HPV). We report a reduction in the number of circulating plasmacytoid DCs (pDCs) in WHIM patients, whereas that of conventional DCs is preserved. This pattern was reproduced in an original mouse model of WS, enabling us to show that the circulating pDC defect can be corrected upon CXCR4 blockade and that pDC differentiation and function are preserved, despite CXCR4 dysfunction. We further identified proper CXCR4 signaling as a critical checkpoint for Langerhans-cell and DC migration from the skin to lymph nodes, with corollary alterations of their activation state and tissue inflammation in a model of HPV-induced dysplasia. Beyond providing new hypotheses to explain the susceptibility of WHIM patients to HPV pathogenesis, this study shows that proper CXCR4 signaling establishes a migration threshold that controls DC egress from CXCL12-containing environments and highlights the critical and subset-specific contribution of CXCR4 signal termination to DC biology.

2.
Cells ; 10(1)2021 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-33466410

RESUMO

Although G protein-coupled receptor kinases (GRKs) have long been known to regulate G protein-coupled receptor (GPCR) desensitization, their more recently characterized functions as scaffolds and signalling adapters underscore that this small family of proteins governs a larger array of physiological functions than originally suspected. This review explores how GRKs contribute to the complex signalling networks involved in the migration of immune cells along chemokine gradients sensed by cell surface GPCRs. We outline emerging evidence indicating that the coordinated docking of several GRKs on an active chemokine receptor determines a specific receptor phosphorylation barcode that will translate into distinct signalling and migration outcomes. The guidance cues for neutrophil migration are emphasized based on several alterations affecting GRKs or GPCRs reported to be involved in pathological conditions.

3.
Artigo em Inglês | MEDLINE | ID: mdl-32956018

RESUMO

Elevated expression of the chemokine receptors CXCR4 and ACKR3 and of their cognate ligand CXCL12 is detected in a wide range of tumors and the tumor microenvironment (TME). Yet, the molecular mechanisms by which the CXCL12/CXCR4/ACKR3 axis contributes to the pathogenesis are complex and not fully understood. To dissect the role of this axis in cancer, we discuss its ability to impinge on canonical and less conventional signaling networks in different cancer cell types; its bidirectional crosstalk, notably with receptor tyrosine kinase (RTK) and other factors present in the TME; and the infiltration of immune cells that support tumor progression. We discuss current and emerging avenues that target the CXCL12/CXCR4/ACKR3 axis. Coordinately targeting both RTKs and CXCR4/ACKR3 and/or CXCL12 is an attractive approach to consider in multitargeted cancer therapies. In addition, inhibiting infiltrating immune cells or reactivating the immune system along with modulating the CXCL12/CXCR4/ACKR3 axis in the TME has therapeutic promise. Expected final online publication date for the Annual Review of Pharmacology and Toxicology, Volume 61 is January 7, 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

4.
Nat Commun ; 11(1): 4855, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32978390

RESUMO

The atypical chemokine receptor 3 (ACKR3) plays a pivotal role in directing the migration of various cellular populations and its over-expression in tumors promotes cell proliferation and invasiveness. The intracellular signaling pathways transducing ACKR3-dependent effects remain poorly characterized, an issue we addressed by identifying the interactome of ACKR3. Here, we report that recombinant ACKR3 expressed in HEK293T cells recruits the gap junction protein Connexin 43 (Cx43). Cx43 and ACKR3 are co-expressed in mouse brain astrocytes and human glioblastoma cells and form a complex in embryonic mouse brain. Functional in vitro studies show enhanced ACKR3 interaction with Cx43 upon ACKR3 agonist stimulation. Furthermore, ACKR3 activation promotes ß-arrestin2- and dynamin-dependent Cx43 internalization to inhibit gap junctional intercellular communication in primary astrocytes. These results demonstrate a functional link between ACKR3 and gap junctions that might be of pathophysiological relevance.


Assuntos
Astrócitos/metabolismo , Comunicação Celular/fisiologia , Conexina 43/metabolismo , Junções Comunicantes/patologia , Receptores CXCR/metabolismo , Animais , Proliferação de Células , Conexina 43/efeitos dos fármacos , Conexinas/metabolismo , Técnicas de Introdução de Genes , Glioblastoma/metabolismo , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Domínios e Motivos de Interação entre Proteínas , Receptores CXCR/agonistas , Receptores CXCR/genética , Transdução de Sinais/fisiologia
5.
J Leukoc Biol ; 107(6): 1123-1135, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32374043

RESUMO

Chemokines play critical roles in numerous physiologic and pathologic processes through their action on seven-transmembrane (TM) receptors. The N-terminal domain of chemokines, which is a key determinant of signaling via its binding within a pocket formed by receptors' TM helices, can be the target of proteolytic processing. An illustrative case of this regulatory mechanism is the natural processing of CXCL12 that generates chemokine variants lacking the first two N-terminal residues. Whereas such truncated variants behave as antagonists of CXCR4, the canonical G protein-coupled receptor of CXCL12, they are agonists of the atypical chemokine receptor 3 (ACKR3/CXCR7), suggesting the implication of different structural determinants in the complexes formed between CXCL12 and its two receptors. Recent analyses have suggested that the CXCL12 N-terminus first engages the TM helices of ACKR3 followed by the receptor N-terminus wrapping around the chemokine core. Here we investigated the first stage of ACKR3-CXCL12 interactions by comparing the activity of substituted or N-terminally truncated variants of CXCL12 toward CXCR4 and ACKR3. We showed that modification of the first two N-terminal residues of the chemokine (K1R or P2G) does not alter the ability of CXCL12 to activate ACKR3. Our results also identified the K1R variant as a G protein-biased agonist of CXCR4. Comparative molecular dynamics simulations of the complexes formed by ACKR3 either with CXCL12 or with the P2G variant identified interactions between the N-terminal 2-4 residues of CXCL12 and a pocket formed by receptor's TM helices 2, 6, and 7 as critical determinants for ACKR3 activation.


Assuntos
Quimiocina CXCL12/química , AMP Cíclico/química , Receptores CXCR4/química , Receptores CXCR/química , Sequência de Aminoácidos , Sítios de Ligação , Quimiocina CXCL11/química , Quimiocina CXCL11/genética , Quimiocina CXCL11/metabolismo , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , AMP Cíclico/metabolismo , Expressão Gênica , Células HEK293 , Compostos Heterocíclicos/química , Compostos Heterocíclicos/farmacologia , Humanos , Simulação de Dinâmica Molecular , Mutação , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Receptores CXCR/genética , Receptores CXCR/metabolismo , Receptores CXCR4/antagonistas & inibidores , Receptores CXCR4/genética , Receptores CXCR4/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , beta-Arrestinas/genética , beta-Arrestinas/metabolismo
7.
Papillomavirus Res ; 8: 100169, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31283993

RESUMO

The linear reverse blotting assays are valid methods for accurate human papillomavirus (HPV) typing required to manage women at risk of developing cervical cancer. However, some samples showed a positive signal in HPV lines but failed to display a positive signal in subsequent typing lines (designated as HPV-X), which indicate that certain types were not available on the respective typing blots. The aim of this study is to elucidate the types or variants of HPV through the high-throughput sequencing (HTS) of 54 ASCUS cervical samples in which the viruses remained untypeable with INNO LiPA HPV® assays. Low-risk (LR)-HPV types (HPV6, 30, 42, 62, 67, 72, 74, 81, 83, 84, 87, 89, 90 and 114), high-risk (HR)-HPV35 and possibly (p)HR-HPV73 were detected among HPV-X. Individual multiple infections (two to seven types) were detected in 40.7% of samples. Twenty-two specimens contained variants characterised by 2-10 changes. HPV30 reached the maximal number of 17 variants with relative abundance inferior or equal to 2.7%. The presence of L1 quasispecies explains why linear reverse blotting assays fail when variants compete or do not match the specific probes. Further studies are needed to measure the LR-HPV quasispecies dynamics and its role during persistent infection.


Assuntos
Colo do Útero/virologia , Variação Genética , Sequenciamento de Nucleotídeos em Larga Escala , Papillomaviridae/genética , Infecções por Papillomavirus/virologia , Quase-Espécies/genética , Sequência de Bases , DNA Viral , Feminino , Genótipo , Humanos , Tipagem Molecular , Papillomaviridae/classificação , Infecções por Papillomavirus/complicações , Filogenia , Análise de Sequência de DNA , Neoplasias do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/etiologia
8.
Mol Pharmacol ; 96(6): 809-818, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31040166

RESUMO

Atypical chemokine receptor 3 (ACKR3), previously known as C-X-C chemokine receptor type 7 (CXCR7), has emerged as a key player in several biologic processes, particularly during development. Its CXCL11 and CXCL12 scavenging activity and atypical signaling properties, together with a new array of other nonchemokine ligands, have established ACKR3 as a main regulator of physiologic processes at steady state and during inflammation. Here, we present a comprehensive review of ACKR3 expression in mammalian tissues in search of a possible connection with the receptor function. Besides the reported roles of ACKR3 during development, we discuss the potential contribution of ACKR3 to the function of the immune system, focusing on the myeloid lineage.


Assuntos
Imunidade Celular/fisiologia , Neutrófilos/imunologia , Neutrófilos/metabolismo , Receptores CXCR/imunologia , Receptores CXCR/metabolismo , Animais , Expressão Gênica , Humanos , Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Receptores CXCR/genética
9.
Immunity ; 50(2): 390-402.e10, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30709741

RESUMO

Neutrophils eliminate pathogens efficiently but can inflict severe damage to the host if they over-activate within blood vessels. It is unclear how immunity solves the dilemma of mounting an efficient anti-microbial defense while preserving vascular health. Here, we identify a neutrophil-intrinsic program that enabled both. The gene Bmal1 regulated expression of the chemokine CXCL2 to induce chemokine receptor CXCR2-dependent diurnal changes in the transcriptional and migratory properties of circulating neutrophils. These diurnal alterations, referred to as neutrophil aging, were antagonized by CXCR4 (C-X-C chemokine receptor type 4) and regulated the outer topology of neutrophils to favor homeostatic egress from blood vessels at night, resulting in boosted anti-microbial activity in tissues. Mice engineered for constitutive neutrophil aging became resistant to infection, but the persistence of intravascular aged neutrophils predisposed them to thrombo-inflammation and death. Thus, diurnal compartmentalization of neutrophils, driven by an internal timer, coordinates immune defense and vascular protection.


Assuntos
Vasos Sanguíneos/imunologia , Ritmo Circadiano/imunologia , Neutrófilos/imunologia , Fagocitose/imunologia , Animais , Vasos Sanguíneos/metabolismo , Candida albicans/imunologia , Candida albicans/fisiologia , Células Cultivadas , Senescência Celular/imunologia , Quimiocina CXCL2/imunologia , Quimiocina CXCL2/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infiltração de Neutrófilos/imunologia , Neutrófilos/metabolismo , Neutrófilos/microbiologia , Receptores CXCR4/imunologia , Receptores CXCR4/metabolismo , Fatores de Tempo
10.
Cytometry A ; 93(7): 681-684, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-30080305

RESUMO

Calcium (Ca2+ ) signaling controls T-cell activation and functions. Ca2+ concentrations are locally detected and controlled by Ca2+ -sensors (STIM1 and 2 detecting the depletion from ER stores channels) and Ca2+ -channels (ORAI1-3 in the cell membrane and VDAC1 in the outer mitochondrial membrane). We first validated and titrated antibodies to assess the expression of these Ca2+ -sensors and -channels in human and murine cells, and further devised a 18-antibodies mass cytometry panel to characterize their expression in primary murine lymphocyte subsets.


Assuntos
Canais de Cálcio/isolamento & purificação , Citometria de Fluxo/métodos , Regulação da Expressão Gênica/genética , Animais , Canais de Cálcio/genética , Membrana Celular/genética , Membrana Celular/metabolismo , Humanos , Subpopulações de Linfócitos/imunologia , Subpopulações de Linfócitos/metabolismo , Camundongos , Membranas Mitocondriais/metabolismo , Proteína ORAI1/genética , Proteína ORAI1/isolamento & purificação , Molécula 1 de Interação Estromal/genética , Molécula 1 de Interação Estromal/isolamento & purificação , Molécula 2 de Interação Estromal/genética , Molécula 2 de Interação Estromal/isolamento & purificação , Canal de Ânion 1 Dependente de Voltagem/genética
11.
Haematologica ; 103(8): 1278-1287, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29724903

RESUMO

Heterozygous germline GATA2 mutations strongly predispose to leukemia, immunodeficiency, and/or lymphoedema. We describe a series of 79 patients (53 families) diagnosed since 2011, made up of all patients in France and Belgium, with a follow up of 2249 patients/years. Median age at first clinical symptoms was 18.6 years (range, 0-61 years). Severe infectious diseases (mycobacteria, fungus, and human papilloma virus) and hematologic malignancies were the most common first manifestations. The probability of remaining symptom-free was 8% at 40 years old. Among the 53 probands, 24 had missense mutations including 4 recurrent alleles, 21 had nonsense or frameshift mutations, 4 had a whole-gene deletion, 2 had splice defects, and 2 patients had complex mutations. There were significantly more cases of leukemia in patients with missense mutations (n=14 of 34) than in patients with nonsense or frameshift mutations (n=2 of 28). We also identify new features of the disease: acute lymphoblastic leukemia, juvenile myelomonocytic leukemia, fatal progressive multifocal leukoencephalopathy related to the JC virus, and immune/inflammatory diseases. A revised International Prognostic Scoring System (IPSS) score allowed a distinction to be made between a stable disease and hematologic transformation. Chemotherapy is of limited efficacy, and has a high toxicity with severe infectious complications. As the mortality rate is high in our cohort (up to 35% at the age of 40), hematopoietic stem cell transplantation (HSCT) remains the best choice of treatment to avoid severe infectious and/or hematologic complications. The timing of HSCT remains difficult to determine, but the earlier it is performed, the better the outcome.


Assuntos
Deficiência de GATA2/epidemiologia , Mutação em Linhagem Germinativa , Adulto Jovem , Adolescente , Adulto , Bélgica , Criança , Pré-Escolar , França , Deficiência de GATA2/complicações , Deficiência de GATA2/genética , Deficiência de GATA2/terapia , Neoplasias Hematológicas/etiologia , Transplante de Células-Tronco Hematopoéticas/métodos , Humanos , Lactente , Recém-Nascido , Infecções/etiologia , Pessoa de Meia-Idade , Mortalidade , Prognóstico , Inquéritos e Questionários
12.
Immunity ; 48(2): 364-379.e8, 2018 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-29466759

RESUMO

Neutrophils are specialized innate cells that require constant replenishment from proliferative bone marrow (BM) precursors as a result of their short half-life. Although it is established that neutrophils are derived from the granulocyte-macrophage progenitor (GMP), the differentiation pathways from GMP to functional mature neutrophils are poorly defined. Using mass cytometry (CyTOF) and cell-cycle-based analysis, we identified three neutrophil subsets within the BM: a committed proliferative neutrophil precursor (preNeu) which differentiates into non-proliferating immature neutrophils and mature neutrophils. Transcriptomic profiling and functional analysis revealed that preNeu require the C/EBPε transcription factor for their generation from the GMP, and their proliferative program is substituted by a gain of migratory and effector function as they mature. preNeus expand under microbial and tumoral stress, and immature neutrophils are recruited to the periphery of tumor-bearing mice. In summary, our study identifies specialized BM granulocytic populations that ensure supply under homeostasis and stress responses.


Assuntos
Células da Medula Óssea/fisiologia , Neutrófilos/fisiologia , Animais , Células da Medula Óssea/imunologia , Proteínas Estimuladoras de Ligação a CCAAT/fisiologia , Linhagem da Célula , Movimento Celular , Proliferação de Células , Células Cultivadas , Perfilação da Expressão Gênica , Humanos , Camundongos , Neoplasias Experimentais/imunologia , Neutrófilos/imunologia
14.
J Pharmacol Exp Ther ; 363(1): 35-44, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28768817

RESUMO

WHIM syndrome is a rare congenital immunodeficiency disease, named after its main clinical manifestations: warts, hypogammaglobulinemia, infections, and myelokathexis, which refers to abnormal accumulation of mature neutrophils in the bone marrow. The disease is primarily caused by C-terminal truncation mutations of the chemokine receptor CXCR4, giving these CXCR4-WHIM mutants a gain of function in response to their ligand CXCL12. Considering the broad functions of CXCR4 in maintaining leukocyte homeostasis, patients are panleukopenic and display altered immune responses, likely as a consequence of impairment in the differentiation and trafficking of leukocytes. Treatment of WHIM patients currently consists of symptom relief, leading to unsatisfactory clinical responses. As an alternative and potentially more effective approach, we tested the potency and efficacy of CXCR4-specific nanobodies on inhibiting CXCR4-WHIM mutants. Nanobodies are therapeutic proteins based on the smallest functional fragments of heavy chain antibodies. They combine the advantages of small-molecule drugs and antibody-based therapeutics due to their relative small size, high stability, and high affinity. We compared the potential of monovalent and bivalent CXCR4-specific nanobodies to inhibit CXCL12-induced CXCR4-WHIM-mediated signaling with the small-molecule clinical candidate AMD3100. The CXCR4-targeting nanobodies displace CXCL12 binding and bind CXCR4-wild type and CXCR4-WHIM (R334X/S338X) mutants and with (sub-) nanomolar affinities. The nanobodies' epitope was mapped to extracellular loop 2 of CXCR4, overlapping with the binding site of CXCL12. Monovalent, and in particular bivalent, nanobodies were more potent than AMD3100 in reducing CXCL12-mediated G protein activation. In addition, CXCR4-WHIM-dependent calcium flux and wound healing of human papillomavirus-immortalized cell lines in response to CXCL12 was effectively inhibited by the nanobodies. Based on these in vitro results, we conclude that CXCR4 nanobodies hold significant potential as alternative therapeutics for CXCR4-associated diseases such as WHIM syndrome.


Assuntos
Especificidade de Anticorpos , Síndromes de Imunodeficiência/imunologia , Síndromes de Imunodeficiência/terapia , Receptores CXCR4/imunologia , Anticorpos de Cadeia Única/imunologia , Anticorpos de Cadeia Única/uso terapêutico , Verrugas/imunologia , Verrugas/terapia , Células HEK293 , Humanos , Síndromes de Imunodeficiência/genética , Mutação , Doenças da Imunodeficiência Primária , Receptores CXCR4/genética , Verrugas/genética
15.
J Exp Med ; 214(7): 2023-2040, 2017 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-28550161

RESUMO

The CXCL12/CXCR4 signaling exerts a dominant role in promoting hematopoietic stem and progenitor cell (HSPC) retention and quiescence in bone marrow. Gain-of-function CXCR4 mutations that affect homologous desensitization of the receptor have been reported in the WHIM Syndrome (WS), a rare immunodeficiency characterized by lymphopenia. The mechanisms underpinning this remain obscure. Using a mouse model with a naturally occurring WS-linked gain-of-function Cxcr4 mutation, we explored the possibility that the lymphopenia in WS arises from defects at the HSPC level. We reported that Cxcr4 desensitization is required for quiescence/cycling balance of murine short-term hematopoietic stem cells and their differentiation into multipotent and downstream lymphoid-biased progenitors. Alteration in Cxcr4 desensitization resulted in decrease of circulating HSPCs in five patients with WS. This was also evidenced in WS mice and mirrored by accumulation of HSPCs in the spleen, where we observed enhanced extramedullary hematopoiesis. Therefore, efficient Cxcr4 desensitization is critical for lymphoid differentiation of HSPCs, and its impairment is a key mechanism underpinning the lymphopenia observed in mice and likely in WS patients.


Assuntos
Diferenciação Celular/genética , Células-Tronco Hematopoéticas/metabolismo , Linfócitos/metabolismo , Receptores CXCR4/genética , Adulto , Animais , Células da Medula Óssea/metabolismo , Transplante de Medula Óssea/métodos , Sobrevivência Celular/genética , Criança , Citometria de Fluxo , Expressão Gênica , Humanos , Síndromes de Imunodeficiência/genética , Síndromes de Imunodeficiência/metabolismo , Contagem de Linfócitos , Camundongos Transgênicos , Mutação , Doenças da Imunodeficiência Primária , Receptores CXCR4/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genética , Baço/citologia , Baço/metabolismo , Verrugas/genética , Verrugas/metabolismo
16.
Nat Commun ; 8: 14253, 2017 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-28181493

RESUMO

Plasmacytoid dendritic cells (pDC) are specialized in secretion of type I interferon in response to pathogens. Here we show that natural monoamines and synthetic amines inhibit pDC activation by RNA viruses. Furthermore, a synthetic analogue of histamine reduces type I interferon production in a mouse model of influenza infection. We identify CXC chemokine receptor 4 (CXCR4) as a receptor used by amines to inhibit pDC. Our study establishes a functional link between natural amines and the innate immune system and identifies CXCR4 as a potential 'on-off' switch of pDC activity with therapeutic potential.


Assuntos
Aminas/farmacologia , Células Dendríticas/metabolismo , Receptores CXCR4/metabolismo , Compostos de Amônio/química , Animais , Células Dendríticas/efeitos dos fármacos , HIV/efeitos dos fármacos , HIV/fisiologia , Histamina/química , Histamina/farmacologia , Humanos , Imidazóis/farmacologia , Interferon Tipo I/metabolismo , Camundongos , Orthomyxoviridae/fisiologia , Receptores Histamínicos/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/farmacologia , Tioureia/análogos & derivados , Tioureia/farmacologia
17.
J Biomol Struct Dyn ; 35(2): 399-412, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26813575

RESUMO

Despite the recent resolutions of the crystal structure of the chemokine receptor CXCR4 in complex with small antagonists or viral chemokine, a description at the molecular level of the interactions between the full-length CXCR4 and its endogenous ligand, the chemokine CXCL12, in relationship with the receptor recognition and activation, is not yet completely elucidated. Moreover, since CXCR4 is able to form dimers, the question of whether the CXCR4-CXCL12 complex has a 1:1 or 2:1 preferential stoichiometry is still an open question. We present here results of coarse-grained protein-protein docking and molecular dynamics simulations of CXCL12 in association with CXCR4 in monomeric and dimeric states. Our proposed models for the 1:1 and 2:1 CXCR4-CXCL12 quaternary structures are consistent with recognition and activation motifs of both partners provided by the available site-directed mutagenesis data. Notably, we observed that in the 2:1 complex, the chemokine N-terminus makes more steady contacts with the receptor residues critical for binding and activation than in the 1:1 structure, suggesting that the 2:1 stoichiometry would favor the receptor signaling activity with respect to the 1:1 association.


Assuntos
Quimiocina CXCL12/química , Conformação Molecular , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Multimerização Proteica , Receptores CXCR4/química , Sítios de Ligação , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , Ligantes , Mutagênese Sítio-Dirigida , Ligação Proteica , Conformação Proteica , Receptores CXCR4/genética , Receptores CXCR4/metabolismo , Relação Estrutura-Atividade
18.
PLoS Pathog ; 12(12): e1006039, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27918748

RESUMO

The productive human papillomavirus (HPV) life cycle is tightly linked to the differentiation and cycling of keratinocytes. Deregulation of these processes and stimulation of cell proliferation by the action of viral oncoproteins and host cell factors underlies HPV-mediated carcinogenesis. Severe HPV infections characterize the wart, hypogammaglobulinemia, infection, and myelokathexis (WHIM) immunodeficiency syndrome, which is caused by gain-of-function mutations in the CXCR4 receptor for the CXCL12 chemokine, one of which is CXCR41013. We investigated whether CXCR41013 interferes in the HPV18 life cycle in epithelial organotypic cultures. Expression of CXCR41013 promoted stabilization of HPV oncoproteins, thus disturbing cell cycle progression and proliferation at the expense of the ordered expression of the viral genes required for virus production. Conversely, blocking CXCR41013 function restored virus production and limited HPV-induced carcinogenesis. Thus, CXCR4 and its potential activation by genetic alterations in the course of the carcinogenic process can be considered as an important host factor for HPV carcinogenesis.


Assuntos
Transformação Celular Viral/fisiologia , Quimiocina CXCL12/metabolismo , Infecções por Papillomavirus/genética , Receptores CXCR4/genética , Transdução de Sinais , Neoplasias Cutâneas/virologia , Animais , Western Blotting , Linhagem Celular , Quimiocina CXCL12/genética , Predisposição Genética para Doença/genética , Xenoenxertos , Papillomavirus Humano 18 , Queratinócitos/metabolismo , Queratinócitos/virologia , Camundongos , Camundongos Nus , Receptores CXCR4/metabolismo , Transdução de Sinais/fisiologia , Neoplasias Cutâneas/genética
19.
J Immunol ; 197(11): 4247-4256, 2016 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-27793999

RESUMO

Ag sampling is a key process in dendritic cell (DC) biology. DCs use constitutive macropinocytosis, receptor-mediated endocytosis, and phagocytosis to capture exogenous Ags for presentation to T cells. We investigated the mechanisms that regulate Ag uptake by DCs in the steady-state and after a short-term LPS exposure in vitro and in vivo. We show that the glucocorticoid-induced leucine zipper protein (GILZ), already known to regulate effector versus regulatory T cell activation by DCs, selectively limits macropinocytosis, but not receptor-mediated phagocytosis, in immature and recently activated DCs. In vivo, the GILZ-mediated inhibition of Ag uptake is restricted to the CD8α+ DC subset, which expresses the highest GILZ level among splenic DC subsets. In recently activated DCs, we further establish that GILZ limits p38 MAPK phosphorylation, providing a possible mechanism for GILZ-mediated macropinocytosis control. Finally, our results demonstrate that the modulation of Ag uptake by GILZ does not result in altered Ag presentation to CD4 T cells but impacts the efficiency of cross-presentation to CD8 T cells. Altogether, our results identify GILZ as an endogenous inhibitor of macropinocytosis in DCs, the action of which contributes to the fine-tuning of Ag cross-presentation.


Assuntos
Antígenos/imunologia , Células Dendríticas/imunologia , Pinocitose/imunologia , Fatores de Transcrição/imunologia , Animais , Apresentação do Antígeno , Antígenos/genética , Linfócitos T CD8-Positivos/imunologia , Camundongos , Camundongos Transgênicos , Pinocitose/genética , Linfócitos T Reguladores/imunologia , Fatores de Transcrição/genética
20.
J Exp Med ; 213(11): 2293-2314, 2016 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-27811056

RESUMO

It is well established that Ly6Chi monocytes develop from common monocyte progenitors (cMoPs) and reside in the bone marrow (BM) until they are mobilized into the circulation. In our study, we found that BM Ly6Chi monocytes are not a homogenous population, as current data would suggest. Using computational analysis approaches to interpret multidimensional datasets, we demonstrate that BM Ly6Chi monocytes consist of two distinct subpopulations (CXCR4hi and CXCR4lo subpopulations) in both mice and humans. Transcriptome studies and in vivo assays revealed functional differences between the two subpopulations. Notably, the CXCR4hi subset proliferates and is immobilized in the BM for the replenishment of functionally mature CXCR4lo monocytes. We propose that the CXCR4hi subset represents a transitional premonocyte population, and that this sequential step of maturation from cMoPs serves to maintain a stable pool of BM monocytes. Additionally, reduced CXCR4 expression on monocytes, upon their exit into the circulation, does not reflect its diminished role in monocyte biology. Specifically, CXCR4 regulates monocyte peripheral cellular activities by governing their circadian oscillations and pulmonary margination, which contributes toward lung injury and sepsis mortality. Together, our study demonstrates the multifaceted role of CXCR4 in defining BM monocyte heterogeneity and in regulating their function in peripheral tissues.


Assuntos
Células da Medula Óssea/citologia , Diferenciação Celular , Monócitos/citologia , Receptores CXCR4/metabolismo , Animais , Antígenos Ly/metabolismo , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Ritmo Circadiano/genética , Endotoxinas/toxicidade , Feminino , Perfilação da Expressão Gênica , Pulmão/irrigação sanguínea , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Camundongos Endogâmicos C57BL , Monócitos/efeitos dos fármacos , Monócitos/metabolismo
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