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1.
PLoS One ; 15(9): e0238347, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32870938

RESUMO

Highly exposed seronegative (HESN) individuals present a unique setting to study mechanisms of protection against HIV acquisition. As natural killer (NK) cell activation and function have been implicated as a correlate of protection in HESN individuals, we sought to better understand the features of NK cells that may confer protection. We used mass cytometry to phenotypically profile NK cells from a cohort of Beninese sex workers and healthy controls. We found that NK cells from HESN women had increased expression of NKG2A, NKp30 and LILRB1, as well as the Fc receptor CD16, and decreased expression of DNAM-1, CD94, Siglec-7, and NKp44. Using functional assessments of NK cells from healthy donors against autologous HIV-infected CD4+ T cells, we observed that NKp30+ and Siglec-7+ cells had improved functional activity. Further, we found that NK cells from HESN women trended towards increased antibody-dependent cellular cytotoxicity (ADCC) activity; this activity correlated with increased CD16 expression. Overall, we identify features of NK cells in HESN women that may contribute to protection from HIV infection. Follow up studies with larger cohorts are warranted to confirm these findings.


Assuntos
Infecções por HIV/patologia , Células Matadoras Naturais/metabolismo , Adulto , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antígenos de Diferenciação Mielomonocítica/metabolismo , Feminino , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Genótipo , Infecções por HIV/imunologia , Humanos , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Lectinas/metabolismo , Modelos Lineares , Receptor 3 Desencadeador da Citotoxicidade Natural/metabolismo , Fenótipo , Receptores de IgG/genética , Receptores de IgG/metabolismo , Profissionais do Sexo
2.
Nat Commun ; 11(1): 4859, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32978401

RESUMO

Cell death is intrinsically linked with immunity. Disruption of an immune-activated MAPK cascade, consisting of MEKK1, MKK1/2, and MPK4, triggers cell death and autoimmunity through the nucleotide-binding leucine-rich repeat (NLR) protein SUMM2 and the MAPK kinase kinase MEKK2. In this study, we identify a Catharanthus roseus receptor-like kinase 1-like (CrRLK1L), named LETUM2/MEDOS1 (LET2/MDS1), and the glycosylphosphatidylinositol (GPI)-anchored protein LLG1 as regulators of mekk1-mkk1/2-mpk4 cell death. LET2/MDS1 functions additively with LET1, another CrRLK1L, and acts genetically downstream of MEKK2 in regulating SUMM2 activation. LET2/MDS1 complexes with LET1 and promotes LET1 phosphorylation, revealing an intertwined regulation between different CrRLK1Ls. LLG1 interacts with the ectodomain of LET1/2 and mediates LET1/2 transport to the plasma membrane, corroborating its function as a co-receptor of LET1/2 in the mekk1-mkk1/2-mpk4 cell death pathway. Thus, our data suggest that a trimeric complex consisting of two CrRLK1Ls LET1, LET2/MDS1, and a GPI-anchored protein LLG1 that regulates the activation of NLR SUMM2 for initiating cell death and autoimmunity.


Assuntos
Autoimunidade/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas Ligadas por GPI/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Autoimunidade/fisiologia , Proteínas de Transporte/imunologia , Catharanthus/genética , Catharanthus/metabolismo , Morte Celular/genética , Proteínas Ligadas por GPI/genética , Regulação da Expressão Gênica de Plantas , Glicosilfosfatidilinositóis , MAP Quinase Quinase Quinases/genética , Sistema de Sinalização das MAP Quinases , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosforilação , Proteínas de Plantas/imunologia , Plantas Geneticamente Modificadas , Interferência de RNA , Transcriptoma
3.
Nat Commun ; 11(1): 4660, 2020 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-32938908

RESUMO

Intratumor spatial heterogeneity facilitates therapeutic resistance in glioblastoma (GBM). Nonetheless, understanding of GBM heterogeneity is largely limited to the surgically resectable tumor core lesion while the seeds for recurrence reside in the unresectable tumor edge. In this study, stratification of GBM to core and edge demonstrates clinically relevant surgical sequelae. We establish regionally derived models of GBM edge and core that retain their spatial identity in a cell autonomous manner. Upon xenotransplantation, edge-derived cells show a higher capacity for infiltrative growth, while core cells demonstrate core lesions with greater therapy resistance. Investigation of intercellular signaling between these two tumor populations uncovers the paracrine crosstalk from tumor core that promotes malignancy and therapy resistance of edge cells. These phenotypic alterations are initiated by HDAC1 in GBM core cells which subsequently affect edge cells by secreting the soluble form of CD109 protein. Our data reveal the role of intracellular communication between regionally different populations of GBM cells in tumor recurrence.


Assuntos
Antígenos CD/metabolismo , Neoplasias Encefálicas/patologia , Glioblastoma/patologia , Histona Desacetilase 1/metabolismo , Proteínas de Neoplasias/metabolismo , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/mortalidade , Feminino , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/mortalidade , Histona Desacetilase 1/antagonistas & inibidores , Histona Desacetilase 1/genética , Histona Desacetilase 2/genética , Histona Desacetilase 2/metabolismo , Humanos , Camundongos SCID , Fenilbutiratos/farmacologia , Transdução de Sinais , Microambiente Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Exp Cell Res ; 395(2): 112204, 2020 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-32735892

RESUMO

BACKGROUND: SARS-CoV2, the agent responsible for the current pandemic, is also causing respiratory distress syndrome (RDS), hyperinflammation and high mortality. It is critical to dissect the pathogenetic mechanisms in order to reach a targeted therapeutic approach. METHODS: In the present investigation, we evaluated the effects of SARS-CoV2 on human bronchial epithelial cells (HBEC). We used RNA-seq datasets available online for identifying SARS-CoV2 potential genes target on human bronchial epithelial cells. RNA expression levels and potential cellular gene pathways have been analyzed. In order to identify possible common strategies among the main pandemic viruses, such as SARS-CoV2, SARS-CoV1, MERS-CoV, and H1N1, we carried out a hypergeometric test of the main genes transcribed in the cells of the respiratory tract exposed to these viruses. RESULTS: The analysis showed that two mechanisms are highly regulated in HBEC: the innate immunity recruitment and the disassembly of cilia and cytoskeletal structure. The granulocyte colony-stimulating factor (CSF3) and dynein heavy chain 7, axonemal (DNAH7) represented respectively the most upregulated and downregulated genes belonging to the two mechanisms highlighted above. Furthermore, the carcinoembryonic antigen-related cell adhesion molecule 7 (CEACAM7) that codifies for a surface protein is highly specific of SARS-CoV2 and not for SARS-CoV1, MERS-CoV, and H1N1, suggesting a potential role in viral entry. In order to identify potential new drugs, using a machine learning approach, we highlighted Flunisolide, Thalidomide, Lenalidomide, Desoximetasone, xylazine, and salmeterol as potential drugs against SARS-CoV2 infection. CONCLUSIONS: Overall, lung involvement and RDS could be generated by the activation and down regulation of diverse gene pathway involving respiratory cilia and muscle contraction, apoptotic phenomena, matrix destructuration, collagen deposition, neutrophil and macrophages recruitment.


Assuntos
Brônquios/metabolismo , Infecções por Coronavirus/genética , Redes Reguladoras de Genes , Pneumonia Viral/genética , Mucosa Respiratória/metabolismo , Transcriptoma , Brônquios/patologia , Antígeno Carcinoembrionário/genética , Antígeno Carcinoembrionário/metabolismo , Infecções por Coronavirus/metabolismo , Descoberta de Drogas/métodos , Dineínas/genética , Dineínas/metabolismo , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Fator Estimulador de Colônias de Granulócitos/genética , Fator Estimulador de Colônias de Granulócitos/metabolismo , Humanos , Imunidade Inata , Aprendizado de Máquina , Pandemias , Pneumonia Viral/metabolismo , Regulação para Cima
5.
Mol Cell Biol ; 40(21)2020 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-32839292

RESUMO

Differentiation status of tumors is correlated with metastatic potential and malignancy. FOXA1 (forkhead box A1) is a transcription factor known to regulate differentiation in certain tissues. Here, we investigate FOXA1 function in human colorectal cancer (CRC). We found that FOXA1 is robustly expressed in the normal human colon but significantly downregulated in colon adenocarcinoma. Applying FOXA1 chromatin immunoprecipitation coupled with deep sequencing and transcriptome analysis upon FOXA1 knockdown in well-differentiated CRC cells and FOXA1 overexpression in poorly differentiated CRC cells, we identified novel protein-coding and lncRNA genes regulated by FOXA1. Among the numerous novel FOXA1 targets we identified, we focused on CEACAM5, a tumor marker and facilitator of cell adhesion. We show that FOXA1 binds to a distal enhancer downstream of CEACAM5 and strongly activates its expression. Consistent with these data, we show that FOXA1 inhibits anoikis in CRC cells. Collectively, our results uncover novel protein-coding and noncoding targets of FOXA1 and suggest a vital role of FOXA1 in enhancing CEACAM5 expression and anoikis resistance in CRC cells.


Assuntos
Neoplasias Colorretais/genética , Redes Reguladoras de Genes , Fator 3-alfa Nuclear de Hepatócito/genética , RNA Longo não Codificante/genética , Adenocarcinoma/genética , Adenocarcinoma/patologia , Anoikis/genética , Antígeno Carcinoembrionário/genética , Antígeno Carcinoembrionário/metabolismo , Diferenciação Celular/genética , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Neoplasias Colorretais/patologia , Elementos Facilitadores Genéticos , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Estudo de Associação Genômica Ampla , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Proteínas/genética , Pseudogenes
6.
Oncogene ; 39(37): 6024-6040, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32778769

RESUMO

Metastatic breast cancer is characterized by high mortality and limited therapeutic target. During tumor metastasis, cytoskeletal reorganization is one of the key steps in the migration and invasion of breast cancer cells. Collapsin response mediator protein 2 (CRMP2) is a cytosolic phosphoprotein that plays an important role in regulating cytoskeletal dynamics. Previous researches have reported that altered CRMP2 expression is associated with breast cancer progression, but the underlying mechanism remains poorly understood. Here, we show that CRMP2 expression is reduced in various subtypes of breast cancers and negatively correlated with lymphatic metastasis. Overexpression of CRMP2 significantly inhibits invasion and stemness in breast cancer cells, while downregulation of CRMP2 promotes cell invasion, which is not required for tubulin polymerization. Mechanistic studies demonstrate that CRMP2 interacts with RECK, prevents RECK degradation, which, in turn, blocks NF-κB and Wnt signaling pathways. Furthermore, we find that phosphorylation of CRMP2 at T514 and S522 remarkably abolishes its functions to bind with RECK and to inhibit cell invasion. Pharmacologic rescue of CRMP2 expression suppressed breast cancer metastasis in vitro and in vivo and stimulated a synergetic effect with FN-1501 that induces CRMP2 dephosphorylation. Collectively, this study highlights the potential of CRMP2 as a therapeutic target in breast cancer metastasis and reveals a distinct mechanism of CRMP2.


Assuntos
Neoplasias da Mama/metabolismo , Proteínas Ligadas por GPI/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Animais , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/etiologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Microtúbulos/metabolismo , Terapia de Alvo Molecular , NF-kappa B/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , Ligação Proteica , Estabilidade Proteica , Via de Sinalização Wnt , Ensaios Antitumorais Modelo de Xenoenxerto
7.
PLoS One ; 15(8): e0225420, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32764749

RESUMO

The H196 residue in SIVmac239 Nef is conserved across the majority of HIV and SIV isolates, lies immediately adjacent to the AP-2 (adaptor protein 2) binding di-leucine domain (ExxxLM195), and is critical for several described AP-2 dependent Nef functions, including the downregulation of tetherin (BST-2/CD317), CD4, and others. Surprisingly, many stocks of the closely related SIVmac251 swarm virus harbor a nef allele encoding a Q196. In SIVmac239, this variant is associated with loss of multiple AP-2 dependent functions. Publicly available sequences for SIVmac251 stocks were mined for variants linked to Q196 that might compensate for functional defects associated with this residue. Variants were engineered into the SIVmac239 backbone and in Nef expression plasmids and flow cytometry was used to examine surface tetherin expression in primary CD4 T cells and surface CD4 expression in SupT1 cells engineered to express rhesus CD4. We found that SIVmac251 stocks that encode a Q196 residue in Nef uniformly also encode an upstream R191 residue. We show that R191 restores the ability of Nef to downregulate tetherin in the presence of Q196 and has a similar but less pronounced impact on CD4 expression. However, a published report showed Q196 commonly evolves to H196 in vivo, suggesting a fitness cost. R191 may represent compensatory evolution to restore the ability to downregulate tetherin lost in viruses harboring Q196.


Assuntos
Antígeno 2 do Estroma da Médula Óssea/metabolismo , Vírus da Imunodeficiência Símia/genética , Proteínas Virais Reguladoras e Acessórias/genética , Animais , Antígenos CD/metabolismo , Antígeno 2 do Estroma da Médula Óssea/genética , Linfócitos T CD4-Positivos/metabolismo , Proteínas Ligadas por GPI/metabolismo , Produtos do Gene nef/metabolismo , Macaca mulatta/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/metabolismo , Vírus da Imunodeficiência Símia/fisiologia , Proteínas Virais Reguladoras e Acessórias/metabolismo
8.
Nat Immunol ; 21(9): 1107-1118, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32788748

RESUMO

In coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, the relationship between disease severity and the host immune response is not fully understood. Here we performed single-cell RNA sequencing in peripheral blood samples of 5 healthy donors and 13 patients with COVID-19, including moderate, severe and convalescent cases. Through determining the transcriptional profiles of immune cells, coupled with assembled T cell receptor and B cell receptor sequences, we analyzed the functional properties of immune cells. Most cell types in patients with COVID-19 showed a strong interferon-α response and an overall acute inflammatory response. Moreover, intensive expansion of highly cytotoxic effector T cell subsets, such as CD4+ effector-GNLY (granulysin), CD8+ effector-GNLY and NKT CD160, was associated with convalescence in moderate patients. In severe patients, the immune landscape featured a deranged interferon response, profound immune exhaustion with skewed T cell receptor repertoire and broad T cell expansion. These findings illustrate the dynamic nature of immune responses during disease progression.


Assuntos
Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Interferon Tipo I/metabolismo , Pneumonia Viral/imunologia , Receptores Imunológicos/metabolismo , Adolescente , Adulto , Idoso , Antígenos CD/genética , Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/genética , Antígenos de Diferenciação de Linfócitos T/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Estudos de Coortes , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/virologia , Feminino , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Proteínas Ligadas por GPI/metabolismo , Humanos , Interferon Tipo I/genética , Interferon Tipo I/imunologia , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/diagnóstico , Pneumonia Viral/virologia , RNA-Seq , Receptores Imunológicos/genética , Receptores Imunológicos/imunologia , Índice de Gravidade de Doença , Análise de Célula Única
9.
Nat Microbiol ; 5(11): 1330-1339, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32704094

RESUMO

Zoonotic coronaviruses (CoVs) are substantial threats to global health, as exemplified by the emergence of two severe acute respiratory syndrome CoVs (SARS-CoV and SARS-CoV-2) and Middle East respiratory syndrome CoV (MERS-CoV) within two decades1-3. Host immune responses to CoVs are complex and regulated in part through antiviral interferons. However, interferon-stimulated gene products that inhibit CoVs are not well characterized4. Here, we show that lymphocyte antigen 6 complex, locus E (LY6E) potently restricts infection by multiple CoVs, including SARS-CoV, SARS-CoV-2 and MERS-CoV. Mechanistic studies revealed that LY6E inhibits CoV entry into cells by interfering with spike protein-mediated membrane fusion. Importantly, mice lacking Ly6e in immune cells were highly susceptible to a murine CoV-mouse hepatitis virus. Exacerbated viral pathogenesis in Ly6e knockout mice was accompanied by loss of hepatic immune cells, higher splenic viral burden and reduction in global antiviral gene pathways. Accordingly, we found that constitutive Ly6e directly protects primary B cells from murine CoV infection. Our results show that LY6E is a critical antiviral immune effector that controls CoV infection and pathogenesis. These findings advance our understanding of immune-mediated control of CoV in vitro and in vivo-knowledge that could help inform strategies to combat infection by emerging CoVs.


Assuntos
Antígenos de Superfície/metabolismo , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Coronavirus/fisiologia , Proteínas Ligadas por GPI/metabolismo , Animais , Antígenos de Superfície/genética , Antígenos de Superfície/imunologia , Betacoronavirus/imunologia , Betacoronavirus/fisiologia , Coronavirus/imunologia , Feminino , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Vírus da SARS/imunologia , Vírus da SARS/fisiologia , Internalização do Vírus
10.
Korean J Parasitol ; 58(3): 249-255, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32615738

RESUMO

Toxoplasma gondii, a ubiquitous, intracellular parasite of the phylum Apicomplexa, infects an estimated one-third of the human population as well as a broad range of warm-blooded animals. We have observed that some tyrosine kinase inhibitors suppressed the growth of T. gondii within host ARPE-10 cells. Among them, afatinib, human epithermal growth factor receptor 2 and 4 (HER2/4) inhibitor, may be used as a therapeutic agent for inhibiting parasite growth with minimal adverse effects on host. In this report, we conducted a proteomic analysis to observe changes in host proteins that were altered via infection with T. gondii and the treatment of HER2/4 inhibitors. Secreting proteins were subjected to a procedure of micor basic reverse phase liquid chromatography, nano-liquid chromatography-mass spectrometry, and ingenuity pathway analysis serially. As a result, the expression level of heterogeneous nuclear ribonucleoprotein K, semaphorin 7A, a GPI membrane anchor, serine/threonine-protein phosphatase 2A, and calpain small subunit 1 proteins were significantly changed, and which were confirmed further by western blot analysis. Changes in various proteins, including these 4 proteins, can be used as a basis for explaining the effects of T. gondii infections and HER2/4 inhibitors.


Assuntos
Afatinib/farmacologia , Afatinib/uso terapêutico , Interações Hospedeiro-Parasita , Proteínas/metabolismo , Receptor ErbB-2/antagonistas & inibidores , Toxoplasma/crescimento & desenvolvimento , Toxoplasmose/tratamento farmacológico , Toxoplasmose/metabolismo , Antígenos CD/metabolismo , Western Blotting , Linhagem Celular , Proteínas Ligadas por GPI/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Humanos , Proteína Fosfatase 2/metabolismo , Proteômica/métodos , Semaforinas/metabolismo
11.
Nat Commun ; 11(1): 3546, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32669559

RESUMO

Advanced ovarian cancer usually spreads to the omentum. However, the omental cell-derived molecular determinants modulating its progression have not been thoroughly characterized. Here, we show that circulating ITLN1 has prognostic significance in patients with advanced ovarian cancer. Further studies demonstrate that ITLN1 suppresses lactotransferrin's effect on ovarian cancer cell invasion potential and proliferation by decreasing MMP1 expression and inducing a metabolic shift in metastatic ovarian cancer cells. Additionally, ovarian cancer-bearing mice treated with ITLN1 demonstrate marked decrease in tumor growth rates. These data suggest that downregulation of mesothelial cell-derived ITLN1 in the omental tumor microenvironment facilitates ovarian cancer progression.


Assuntos
Carcinoma Epitelial do Ovário/secundário , Citocinas/metabolismo , Lectinas/metabolismo , Omento/patologia , Neoplasias Ovarianas/patologia , Neoplasias Peritoneais/secundário , Animais , Carcinoma Epitelial do Ovário/sangue , Carcinoma Epitelial do Ovário/mortalidade , Carcinoma Epitelial do Ovário/terapia , Linhagem Celular Tumoral/transplante , Movimento Celular , Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Citocinas/administração & dosagem , Citocinas/sangue , Modelos Animais de Doenças , Regulação para Baixo , Feminino , Proteínas Ligadas por GPI/administração & dosagem , Proteínas Ligadas por GPI/sangue , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Lactoferrina/metabolismo , Lectinas/administração & dosagem , Lectinas/sangue , Metaloproteinase 1 da Matriz/metabolismo , Camundongos , Invasividade Neoplásica/patologia , Neoplasias Ovarianas/sangue , Neoplasias Ovarianas/mortalidade , Neoplasias Ovarianas/terapia , Ovário , Proteínas Recombinantes/administração & dosagem , Taxa de Sobrevida , Microambiente Tumoral
12.
J Virol ; 94(18)2020 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-32641482

RESUMO

C3A is a subclone of the human hepatoblastoma HepG2 cell line with strong contact inhibition of growth. We fortuitously found that C3A was more susceptible to human coronavirus HCoV-OC43 infection than HepG2, which was attributed to the increased efficiency of virus entry into C3A cells. In an effort to search for the host cellular protein(s) mediating the differential susceptibility of the two cell lines to HCoV-OC43 infection, we found that ArfGAP with dual pleckstrin homology (PH) domains 2 (ADAP2), gamma-interferon-inducible lysosome/endosome-localized thiolreductase (GILT), and lymphocyte antigen 6 family member E (LY6E), the three cellular proteins identified to function in interference with virus entry, were expressed at significantly higher levels in HepG2 cells. Functional analyses revealed that ectopic expression of LY6E, but not GILT or ADAP2, in HEK 293 cells inhibited the entry of HCoV-O43. While overexpression of LY6E in C3A and A549 cells efficiently inhibited the infection of HCoV-OC43, knockdown of LY6E expression in HepG2 significantly increased its susceptibility to HCoV-OC43 infection. Moreover, we found that LY6E also efficiently restricted the entry mediated by the envelope spike proteins of other human coronaviruses, including the currently pandemic SARS-CoV-2. Interestingly, overexpression of serine protease TMPRSS2 or amphotericin treatment significantly neutralized the IFN-inducible transmembrane 3 (IFITM3) restriction of human coronavirus (CoV) entry, but did not compromise the effect of LY6E on the entry of human coronaviruses. The work reported herein thus demonstrates that LY6E is a critical antiviral immune effector that controls CoV infection and pathogenesis via a mechanism distinct from other factors that modulate CoV entry.IMPORTANCE Virus entry into host cells is one of the key determinants of host range and cell tropism and is subjected to the control of host innate and adaptive immune responses. In the last decade, several interferon-inducible cellular proteins, including IFITMs, GILT, ADAP2, 25CH, and LY6E, had been identified to modulate the infectious entry of a variety of viruses. Particularly, LY6E was recently identified as a host factor that facilitates the entry of several human-pathogenic viruses, including human immunodeficiency virus, influenza A virus, and yellow fever virus. Identification of LY6E as a potent restriction factor of coronaviruses expands the biological function of LY6E and sheds new light on the immunopathogenesis of human coronavirus infection.


Assuntos
Antígenos de Superfície/metabolismo , Betacoronavirus/fisiologia , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Coronavirus/fisiologia , Interações Hospedeiro-Patógeno , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , Internalização do Vírus , Sequência de Aminoácidos , Anfotericina B/farmacologia , Betacoronavirus/efeitos dos fármacos , Linhagem Celular , Coronavirus/efeitos dos fármacos , Infecções por Coronavirus/epidemiologia , Suscetibilidade a Doenças , Evolução Molecular , Proteínas Ligadas por GPI/metabolismo , Humanos , Pandemias , Pneumonia Viral/epidemiologia , Sinais Direcionadores de Proteínas , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo
13.
Cells ; 9(8)2020 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-32707842

RESUMO

The ectonucleotidases CD39 and CD73 regulate immune responses by balancing extracellular ATP and adenosine in inflammation and are likely to be involved in the pathophysiology of COVID-19. Here, we analyzed CD39 and CD73 on different lymphocyte populations in a small cohort of COVID-19 patients and in healthy individuals. We describe a significantly lower level of expression of CD73 on cytotoxic lymphocyte populations, including CD8+ T, natural killer T (NKT), and natural killer (NK) cells, during COVID-19. Interestingly, the decrease of CD73 on CD8+ T cells and NKT cells correlated with serum ferritin levels. Furthermore, we observed distinct functional differences between the CD73+ and CD73- subsets of CD8+ T cells and NKT cells with regard to cytokine/toxin secretion. In COVID-19 patients, the majority of the CD73-CD8+ T cells were capable of secreting granzyme B, perforin, tumor necrosis factor (TNF-α) or interferon-gamma (IFN-γ). To conclude, in this first study of CD39 and CD73 expression of lymphocytes in COVID-19, we show that CD8+ T cells and NKT cells lacking CD73 possess a significantly higher cytotoxic effector functionality compared to their CD73+ counterparts. Future studies should investigate differences of cellular CD39 and CD73 expression in patients at different disease stages and their potential as prognostic markers or targets for immunomodulatory therapies.


Assuntos
5'-Nucleotidase/metabolismo , Apirase/metabolismo , Infecções por Coronavirus/imunologia , Células Matadoras Naturais/imunologia , Células T Matadoras Naturais/imunologia , Pneumonia Viral/imunologia , Linfócitos T Citotóxicos/imunologia , Adenosina/metabolismo , Adulto , Idoso , Betacoronavirus , Infecções por Coronavirus/enzimologia , Feminino , Proteínas Ligadas por GPI/metabolismo , Granzimas/metabolismo , Humanos , Inflamação/enzimologia , Inflamação/imunologia , Interferon gama/metabolismo , Masculino , Pessoa de Meia-Idade , Pandemias , Perforina/metabolismo , Pneumonia Viral/enzimologia , Transdução de Sinais/imunologia , Linfócitos T Citotóxicos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
14.
Carcinogenesis ; 41(5): 571-581, 2020 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-32648918

RESUMO

Cripto-1 (CR1), an oncofetal protein, had been implied to reactivate in some cancers. However, the relationship between CR1 expression and patient outcomes and the tumor biological function of CR1 contributing to invasion and metastasis in hepatocellular carcinoma (HCC) is poorly defined. In this study, we demonstrated that CR1 was expressed in over 80% of HCCs in a training cohort (n = 242) and a validation cohort (n = 159). High CR1 expression was significantly correlated with aggressive HCC phenotypes (i.e. portal vein tumor thrombus, microscopic vascular invasion, multiple tumors and poor tumor differentiation). In both the training and validation cohorts, patients with high CR1 expression had remarkably shorter disease-free survival and overall survival rates than those with low CR1 expression. A series in vitro and in vivo assays showed that CR1 substantially promoted HCC cell migration, invasion and metastasis. Mechanistically, we demonstrated that CR1 induced HCC cells to undergo epithelial-mesenchymal transition through activating the Akt/NFκB/p65 signaling. Chromatin immunoprecipitation assay showed that NFκB/p65 enhanced CR1 expression by binding its promoter. Thus, CR1 and NFκB/p65 form a positive feedback loop that sustained the process of migration and invasion of HCC. Therefore, CR1 plays an important role in HCC invasion and metastasis and may be an effective and reliable prognostic biomarker for HCC recurrence after resection. Targeting CR1 may be a promising treatment for HCC.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/patologia , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/secundário , Proteínas de Neoplasias/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/cirurgia , Movimento Celular , Proliferação de Células , Feminino , Proteínas Ligadas por GPI/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/cirurgia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/cirurgia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Prognóstico , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Carcinogenesis ; 41(5): 571-581, 2020 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-32649753

RESUMO

Cripto-1 (CR1), an oncofetal protein, had been implied to reactivate in some cancers. However, the relationship between CR1 expression and patient outcomes and the tumor biological function of CR1 contributing to invasion and metastasis in hepatocellular carcinoma (HCC) is poorly defined. In this study, we demonstrated that CR1 was expressed in over 80% of HCCs in a training cohort (n = 242) and a validation cohort (n = 159). High CR1 expression was significantly correlated with aggressive HCC phenotypes (i.e. portal vein tumor thrombus, microscopic vascular invasion, multiple tumors and poor tumor differentiation). In both the training and validation cohorts, patients with high CR1 expression had remarkably shorter disease-free survival and overall survival rates than those with low CR1 expression. A series in vitro and in vivo assays showed that CR1 substantially promoted HCC cell migration, invasion and metastasis. Mechanistically, we demonstrated that CR1 induced HCC cells to undergo epithelial-mesenchymal transition through activating the Akt/NFκB/p65 signaling. Chromatin immunoprecipitation assay showed that NFκB/p65 enhanced CR1 expression by binding its promoter. Thus, CR1 and NFκB/p65 form a positive feedback loop that sustained the process of migration and invasion of HCC. Therefore, CR1 plays an important role in HCC invasion and metastasis and may be an effective and reliable prognostic biomarker for HCC recurrence after resection. Targeting CR1 may be a promising treatment for HCC.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/patologia , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/secundário , Proteínas de Neoplasias/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/cirurgia , Movimento Celular , Proliferação de Células , Feminino , Proteínas Ligadas por GPI/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/cirurgia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/cirurgia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Prognóstico , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Proc Natl Acad Sci U S A ; 117(26): 15104-15111, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32541044

RESUMO

Five small protein domains, the CC-domains, at the N terminus of the RECK protein, play essential roles in signaling by WNT7A and WNT7B in the context of central nervous system angiogenesis and blood-brain barrier formation and maintenance. We have determined the structure of CC domain 4 (CC4) at 1.65-Å resolution and find that it folds into a compact four-helix bundle with three disulfide bonds. The CC4 structure, together with homology modeling of CC1, reveals the surface locations of critical residues that were shown in previous mutagenesis studies to mediate GPR124 binding and WNT7A/WNT7B recognition and signaling. Surprisingly, sequence and structural homology searches reveal no other cell-surface or secreted domains in vertebrates that resemble the CC domain, a pattern that is in striking contrast to other ancient and similarly sized domains, such as Epidermal Growth Factor, Fibronectin Type 3, Immunoglobulin, and Thrombospondin type 1 domains, which are collectively present in hundreds of proteins.


Assuntos
Evolução Molecular , Proteínas Ligadas por GPI/química , Proteínas Ligadas por GPI/metabolismo , Sequência de Aminoácidos , Animais , Caenorhabditis elegans , Proteínas Ligadas por GPI/genética , Humanos , Camundongos , Domínios Proteicos , Alinhamento de Sequência
17.
Proc Natl Acad Sci U S A ; 117(27): 15620-15631, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32576689

RESUMO

Repulsive guidance molecules (RGMs) are cell surface proteins that regulate the development and homeostasis of many tissues and organs, including the nervous, skeletal, and immune systems. They control fundamental biological processes, such as migration and differentiation by direct interaction with the Neogenin (NEO1) receptor and function as coreceptors for the bone morphogenetic protein (BMP)/growth differentiation factor (GDF) family. We determined crystal structures of all three human RGM family members in complex with GDF5, as well as the ternary NEO1-RGMB-GDF5 assembly. Surprisingly, we show that all three RGMs inhibit GDF5 signaling, which is in stark contrast to RGM-mediated enhancement of signaling observed for other BMPs, like BMP2. Despite their opposite effect on GDF5 signaling, RGMs occupy the BMP type 1 receptor binding site similar to the observed interactions in RGM-BMP2 complexes. In the NEO1-RGMB-GDF5 complex, RGMB physically bridges NEO1 and GDF5, suggesting cross-talk between the GDF5 and NEO1 signaling pathways. Our crystal structures, combined with structure-guided mutagenesis of RGMs and BMP ligands, binding studies, and cellular assays suggest that RGMs inhibit GDF5 signaling by competing with GDF5 type 1 receptors. While our crystal structure analysis and in vitro binding data initially pointed towards a simple competition mechanism between RGMs and type 1 receptors as a possible basis for RGM-mediated GDF5 inhibition, further experiments utilizing BMP2-mimicking GDF5 variants clearly indicate a more complex mechanism that explains how RGMs can act as a functionality-changing switch for two structurally and biochemically similar signaling molecules.


Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Proteínas Ligadas por GPI/metabolismo , Fator 5 de Diferenciação de Crescimento/metabolismo , Proteína da Hemocromatose/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteína Morfogenética Óssea 2/metabolismo , Proteína Morfogenética Óssea 2/ultraestrutura , Moléculas de Adesão Celular Neuronais/ultraestrutura , Cristalografia por Raios X , Proteínas Ligadas por GPI/ultraestrutura , Fator 5 de Diferenciação de Crescimento/ultraestrutura , Proteína da Hemocromatose/ultraestrutura , Proteínas de Membrana/metabolismo , Proteínas de Membrana/ultraestrutura , Proteínas do Tecido Nervoso/ultraestrutura , Domínios Proteicos , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura , Transdução de Sinais
18.
Nat Commun ; 11(1): 3177, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32576819

RESUMO

Vascular permeability and plasma leakage are immune-pathologies of severe dengue virus (DENV) infection, but the mechanisms underlying the exacerbated inflammation during DENV pathogenesis are unclear. Here, we demonstrate that TLR2, together with its co-receptors CD14 and TLR6, is an innate sensor of DENV particles inducing inflammatory cytokine expression and impairing vascular integrity in vitro. Blocking TLR2 prior to DENV infection in vitro abrogates NF-κB activation while CD14 and TLR6 block has a moderate effect. Moreover, TLR2 block prior to DENV infection of peripheral blood mononuclear cells prevents activation of human vascular endothelium, suggesting a potential role of the TLR2-responses in vascular integrity. TLR2 expression on CD14 + + classical monocytes isolated in an acute phase from DENV-infected pediatric patients correlates with severe disease development. Altogether, these data identify a role for TLR2 in DENV infection and provide insights into the complex interaction between the virus and innate receptors that may underlie disease pathogenesis.


Assuntos
Vírus da Dengue/metabolismo , Dengue/imunologia , Monócitos/metabolismo , Receptor 2 Toll-Like/metabolismo , Permeabilidade Capilar , Quimiocinas/metabolismo , Criança , Pré-Escolar , Citocinas/metabolismo , Dengue/virologia , Endotélio Vascular/metabolismo , Feminino , Proteínas Ligadas por GPI/metabolismo , Regulação da Expressão Gênica , Humanos , Imunidade Inata , Inflamação , Leucócitos Mononucleares/metabolismo , Receptores de Lipopolissacarídeos/metabolismo , Masculino , NF-kappa B/metabolismo , Receptores de IgG/metabolismo , Índice de Gravidade de Doença , Receptor 6 Toll-Like
19.
Nat Commun ; 11(1): 3175, 2020 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-32581250

RESUMO

Pancreatic cancer is the fourth leading cause of cancer-related deaths in Japan. To identify risk loci, we perform a meta-analysis of three genome-wide association studies comprising 2,039 pancreatic cancer patients and 32,592 controls in the Japanese population. Here, we identify 3 (13q12.2, 13q22.1, and 16p12.3) genome-wide significant loci (P < 5.0 × 10-8), of which 16p12.3 has not been reported in the Western population. The lead single nucleotide polymorphism (SNP) at 16p12.3 is rs78193826 (odds ratio = 1.46, 95% confidence interval = 1.29-1.66, P = 4.28 × 10-9), an Asian-specific, nonsynonymous glycoprotein 2 (GP2) gene variant. Associations between selected GP2 gene variants and pancreatic cancer are replicated in 10,822 additional cases and controls of East Asian origin. Functional analyses using cell lines provide supporting evidence of the effect of rs78193826 on KRAS activity. These findings suggest that GP2 gene variants are probably associated with pancreatic cancer susceptibility in populations of East Asian ancestry.


Assuntos
Proteínas Ligadas por GPI/genética , Predisposição Genética para Doença/genética , Neoplasias Pancreáticas/genética , Grupo com Ancestrais do Continente Asiático/genética , Linhagem Celular Tumoral , Bases de Dados Genéticas , Proteínas Ligadas por GPI/metabolismo , Loci Gênicos , Pleiotropia Genética , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Polimorfismo de Nucleotídeo Único
20.
BMC Cardiovasc Disord ; 20(1): 214, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32375640

RESUMO

BACKGROUND: Epicardial adipose tissue (EAT) remodeling and adipocytokines are associated with structural remodeling in atrial fibrillation (AF). However, the role of omentin-1, a novel adipocytokine, in structural remodeling remains unknown. METHODS: Hematoxylin and eosin (H&E) and Masson's trichrome stains were used to investigate the histology of EAT and right atrial appendages. The expression levels of adipocytokines in these human samples were determined by immunohistochemical assay and western blotting. Models of transforming growth factor (TGF)-ß1-induced activation of cardiac fibroblasts (CFs) and TGF-ß1-induced endothelial-mesenchymal transition (EndMT) of human umbilical vein endothelial cell (HUVEC) were established to explore roles of omentin-1 in these processes. To determine changes in adipocytokines secretion under hypoxia conditions, adipocytes were treated with 5% O2 and 95% N2, and then CFs and HUVECs were co-cultured with the conditioned medium of adipocytes to determine the effects of hypoxia-treated adipocytes on these cells. RESULTS: Expression of omentin-1 was downregulated in the EAT and right atrial appendages from patients with AF compared to samples from patients without AF, while the TGF-ß1 level was upregulated in EAT from patients with AF. EAT from patients with AF exhibited adipocyte hypertrophy and severe interstitial fibrosis. Omentin-1 inhibited TGF-ß1-induced CF activation and reversed TGF-ß1-induced HUVEC EndMT. Adipocytes treated with hypoxia exhibited downregulation of omentin-1 and partly activated CFs. CONCLUSIONS: This study demonstrated that omentin-1 was an antifibrotic adipocytokine and was downregulated in patients with AF, which was partly mediated by hypoxia.


Assuntos
Adipócitos/metabolismo , Fibrilação Atrial/metabolismo , Citocinas/metabolismo , Doenças das Valvas Cardíacas/metabolismo , Lectinas/metabolismo , Comunicação Parácrina , Células 3T3-L1 , Adipócitos/patologia , Adulto , Idoso , Animais , Fibrilação Atrial/patologia , Fibrilação Atrial/fisiopatologia , Hipóxia Celular , Técnicas de Cocultura , Transição Epitelial-Mesenquimal , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose , Proteínas Ligadas por GPI/metabolismo , Doenças das Valvas Cardíacas/patologia , Doenças das Valvas Cardíacas/fisiopatologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Ratos , Transdução de Sinais , Fator de Crescimento Transformador beta1/metabolismo , Função Ventricular Esquerda , Remodelação Ventricular
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