RESUMO
Peptidyl prolyl cis/trans isomerases (PPIases) are a superfamily of proteins ubiquitously distributed among living organisms, which function primarily to assist the folding and structuring of unfolded and partially folded polypeptide chains and proteins. In this review, we focus specifically on the Macrophage Infectivity Potentiator (MIP)-like PPIases, which are members of the immunophilin family of FK506-binding proteins (FKBP). MIP-like PPIases have accessory roles in virulence and are candidates for inclusion in vaccines protective against both animal and human bacterial pathogens. A structural vaccinology approach obviates any issues over molecular mimicry and potential cross-reactivity with human FKBP proteins and studies with a representative antigen, the Neisseria meningitidis-MIP, support this strategy. Moreover, a dual approach of vaccination and drug targeting could be considered for controlling bacterial infectious diseases of humans and animals.
Assuntos
Proteínas de Bactérias/imunologia , Vacinas Bacterianas/imunologia , Macrófagos/imunologia , Peptidilprolil Isomerase/imunologia , Proteínas de Ligação a Tacrolimo/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Sequência de Aminoácidos , Animais , Infecções Bacterianas/imunologia , Infecções Bacterianas/prevenção & controle , Desenho de Fármacos , Humanos , Legionella pneumophila/imunologia , Meningite Meningocócica/imunologia , Meningite Meningocócica/microbiologia , Meningite Meningocócica/prevenção & controle , Dados de Sequência Molecular , Neisseria meningitidis/imunologia , Neisseria meningitidis/patogenicidadeRESUMO
The unique proline isomerase Pin1 is pivotal for protecting against age-dependent neurodegeneration in Alzheimer's disease (AD), with its inhibition providing a molecular link between tangle and plaque pathologies. Pin1 is oxidatively modified in human AD brains, but little is known about its regulatory mechanisms and pathological significance of such Pin1 modification. In this paper, our determination of crystal structures of oxidized Pin1 reveals a series of Pin1 oxidative modifications on Cys113 in a sequential fashion. Cys113 oxidization is further confirmed by generating antibodies specifically recognizing oxidized Cys113 of Pin1. Furthermore, Pin1 oxidation on Cys113 inactivates its catalytic activity in vitro, and Ala point substitution of Cys113 inactivates the ability of Pin1 to isomerize tau as well as to promote protein turnover of tau and APP. Moreover, redox regulation affects Pin1 subcellular localization and Pin1-mediated neuronal survival in response to hypoxia treatment. Importantly, Cys113-oxidized Pin1 is significantly increased in human AD brain comparing to age-matched controls. These results not only identify a novel Pin1 oxidation site to be the critical catalytic residue Cys113, but also provide a novel oxidative regulation mechanism for inhibiting Pin1 activity in AD. These results suggest that preventing Pin1 oxidization might help to reduce the risk of AD.
Assuntos
Doença de Alzheimer/metabolismo , Peptidilprolil Isomerase/química , Peptidilprolil Isomerase/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Anticorpos , Domínio Catalítico , Linhagem Celular Tumoral , Hipocampo/metabolismo , Humanos , Peptidilprolil Isomerase de Interação com NIMA , Oxirredução , Peptidilprolil Isomerase/imunologia , Proteínas tau/metabolismoRESUMO
Streptococcus gordonii is a commensal gram-positive bacterium that resides in the human oral cavity, and is one of the most common causes of infective endocarditis (IE). Bacterial surface molecules play an important role in establishing IE, and several S. gordonii proteins have been implicated in binding to host cells during the establishment of IE. In this study, we identified a putative lipoprotein, peptidyl-prolyl cis/trans isomerase (PpiA), and clarified its role in evasion of phagocytosis by macrophages. Attenuation of the gene encoding prolipoprotein diacylglyceryl transferase (Lgt) altered the localization of PpiA from the cell surface to the culture supernatant, indicating that PpiA is lipid-anchored in the cell membrane by Lgt. Both human and murine macrophages showed higher phagocytic activity towards ppiA and lgt mutants than the wild-type, indicating that the presence of PpiA suppresses phagocytosis of S. gordonii. Human macrophages treated with dextran sulfate had significantly impaired phagocytosis of S. gordonii, suggesting that class A scavenger receptors in human macrophages are involved in the phagocytosis of S. gordonii. These results provide evidence that S. gordonii lipoprotein PpiA plays an important role in inhibiting phagocytic engulfment and in evasion of the host immune response.
Assuntos
Macrófagos/microbiologia , Peptidilprolil Isomerase/imunologia , Fagocitose/imunologia , Streptococcus gordonii/enzimologia , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Linhagem Celular Tumoral , Células Cultivadas , Sulfato de Dextrana/farmacologia , Inativação Gênica , Humanos , Evasão da Resposta Imune/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/microbiologia , Lipídeos de Membrana/genética , Lipídeos de Membrana/imunologia , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/imunologia , Camundongos , Mutação/genética , Peptidilprolil Isomerase/genética , Fagocitose/efeitos dos fármacos , RNA Interferente Pequeno , Receptores Depuradores Classe A/imunologia , Streptococcus gordonii/genética , Transferases/genética , Transferases/imunologiaRESUMO
This study tested the hypothesis that priming the neutrophil respiratory burst requires both granule exocytosis and activation of the prolyl isomerase Pin1. Fusion proteins containing the TAT cell permeability sequence and either the SNARE domain of syntaxin-4 or the N-terminal SNARE domain of SNAP-23 were used to examine the role of granule subsets in TNF-mediated respiratory burst priming using human neutrophils. Concentration-inhibition curves for exocytosis of individual granule subsets and for priming of fMLF-stimulated superoxide release and phagocytosis-stimulated H2O2 production were generated. Maximal inhibition of priming ranged from 72 to 88%. Linear regression lines for inhibition of priming versus inhibition of exocytosis did not differ from the line of identity for secretory vesicles and gelatinase granules, while the slopes or the y-intercepts were different from the line of identity for specific and azurophilic granules. Inhibition of Pin1 reduced priming by 56%, while exocytosis of secretory vesicles and specific granules was not affected. These findings indicate that exocytosis of secretory vesicles and gelatinase granules and activation of Pin1 are independent events required for TNF-mediated priming of neutrophil respiratory burst.
Assuntos
Exocitose/imunologia , Neutrófilos/imunologia , Peptidilprolil Isomerase/imunologia , Explosão Respiratória/imunologia , Vesículas Secretórias/imunologia , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/imunologia , Exocitose/efeitos dos fármacos , Feminino , Humanos , Masculino , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Peptidilprolil Isomerase de Interação com NIMA , Neutrófilos/enzimologia , Peptidilprolil Isomerase/metabolismo , Proteínas Qa-SNARE/imunologia , Proteínas Qa-SNARE/metabolismo , Proteínas Qb-SNARE/imunologia , Proteínas Qb-SNARE/metabolismo , Proteínas Qc-SNARE/imunologia , Proteínas Qc-SNARE/metabolismo , Explosão Respiratória/efeitos dos fármacos , Vesículas Secretórias/enzimologia , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammation in joints and subsequent destruction of cartilage and bone. Inflammatory mediators such as PGs and proinflammatory cytokines contribute to RA progress. Pin1, a peptidyl prolyl isomerase, plays important pathophysiological roles in several diseases, including cancer and neurodegeneration. We found that both Pin1 and cyclooxygenase-2 (COX-2) were highly expressed in ankle tissues of type II collagen-induced RA mice. HTB-94 cells overexpressing Pin1 and primary cultured human chondrocytes showed increased basal expression of proinflammatory proteins (COX-2, inducible NO synthase, TNF-alpha, and IL-1beta). Site-directed mutagenesis revealed that Pin1-mediated transcriptional activation of COX-2 was coordinately regulated by NF-kappaB, CREB, and C/EBP. Gel shift, reporter gene, and Western blot analyses confirmed that NF-kappaB, CREB, and C/EBP were consistently activated in chondrocytes overexpressing Pin1. Treatment of RA mice with juglone, a chemical inhibitor of Pin1, significantly reduced RA progress and COX-2 expression in the ankle tissues. Moreover, juglone dose dependently decreased the basal COX-2 expression in primary cultured chondrocytes from RA patients. These results demonstrate that Pin1 induction during RA progress stimulates proinflammatory protein expression by activating NF-kappaB, CREB, and C/EBP, and suggest that Pin1 is a potential therapeutic target of RA.
Assuntos
Artrite Reumatoide/imunologia , Condrócitos/imunologia , Naftoquinonas/uso terapêutico , Peptidilprolil Isomerase/imunologia , Animais , Articulação do Tornozelo/efeitos dos fármacos , Articulação do Tornozelo/imunologia , Articulação do Tornozelo/patologia , Artrite Reumatoide/tratamento farmacológico , Linhagem Celular , Células Cultivadas , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Colágeno Tipo II/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/imunologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Ciclo-Oxigenase 2/efeitos dos fármacos , Ciclo-Oxigenase 2/imunologia , Ciclo-Oxigenase 2/metabolismo , Genes Reporter/genética , Genes Reporter/imunologia , Humanos , Interleucina-1beta/imunologia , Interleucina-1beta/metabolismo , Camundongos , NF-kappa B/imunologia , NF-kappa B/metabolismo , Peptidilprolil Isomerase de Interação com NIMA , Naftoquinonas/administração & dosagem , Óxido Nítrico Sintase Tipo II/imunologia , Óxido Nítrico Sintase Tipo II/metabolismo , Peptidilprolil Isomerase/antagonistas & inibidores , Peptidilprolil Isomerase/metabolismo , Esteroide Isomerases/imunologia , Esteroide Isomerases/metabolismo , Transfecção , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Polyclonal antibodies raised against full-length antigens are often used for localization experiments. Exact knowledge of epitopes in the antigen recognized by the antiserum is important if the target antigen belongs to a large family of proteins which are highly conserved. We have shown that epitope mapping using peptide microarrays represents a powerful tool for determination of immunodominat regions in a proteome-wide manner. As examples we show results of epitope mapping using peptide microarrays displaying overlapping peptide scans through either all human cyclophilins or all human FK506-binding proteins.
Assuntos
Anticorpos/imunologia , Mapeamento de Epitopos/métodos , Peptídeos/imunologia , Análise Serial de Proteínas/métodos , Sequência de Aminoácidos , Anticorpos/química , Reações Cruzadas , Ciclofilinas/química , Ciclofilinas/imunologia , Humanos , Dados de Sequência Molecular , Peptídeos/química , Peptidilprolil Isomerase/química , Peptidilprolil Isomerase/imunologia , Estrutura Terciária de Proteína , Alinhamento de Sequência , Proteínas de Ligação a Tacrolimo/química , Proteínas de Ligação a Tacrolimo/imunologiaRESUMO
The mechanisms by which cytokine signals prevent the activation and mitochondrial targeting of the proapoptotic protein Bax are unclear. Here we show, using primary human eosinophils, that in the absence of the prosurvival cytokines granulocyte-macrophage colony-stimulating factor and interleukin 5, Bax spontaneously underwent activation and initiated mitochondrial disruption. Inhibition of Bax resulted in less eosinophil apoptosis, even in the absence of cytokines. Granulocyte-macrophage colony-stimulating factor induced activation of the kinase Erk1/2, which phosphorylated Thr167 of Bax; this facilitated new interaction of Bax with the prolyl isomerase Pin1. Blockade of Pin1 led to cleavage and mitochondrial translocation of Bax and caspase activation, regardless of the presence of cytokines. Our findings indicate that Pin1 is a key mediator of prosurvival signaling and is a regulator of Bax function.
Assuntos
Eosinófilos/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Interleucina-5/imunologia , Peptidilprolil Isomerase/imunologia , Proteína X Associada a bcl-2/imunologia , Morte Celular , Sobrevivência Celular , Células Cultivadas , Eosinófilos/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/imunologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Interleucina-5/metabolismo , Mitocôndrias/imunologia , Mitocôndrias/metabolismo , Peptidilprolil Isomerase de Interação com NIMA , Peptidilprolil Isomerase/metabolismo , Fosforilação , Transporte Proteico , Proteína X Associada a bcl-2/metabolismoRESUMO
Cell growth and differentiation are highly controlled processes mediated by effector molecules, which are regulated by posttranslational chemical modifications. Adaptor molecules are critical players in these mechanisms because of their ability to simultaneously interact with multiple effector molecules and orchestrate the assembly of signaling complexes downstream of activated surface receptors. One family of adaptor molecules includes the CrkII/CrkL proteins that are also involved in the regulation of lymphocyte function. Although Crk proteins are amenable to regulation by protein tyrosine kinases, recent data suggest that peptidyl-prolyl cis-trans isomerases (PPIases) can alter their conformation and hence their ability to associate with binding partners. This emerging new function of PPIases is the subject of the current review.
Assuntos
Linfócitos/enzimologia , Peptidilprolil Isomerase/metabolismo , Proteínas Proto-Oncogênicas c-crk/química , Proteínas Proto-Oncogênicas c-crk/metabolismo , Transdução de Sinais/fisiologia , Animais , Humanos , Linfócitos/imunologia , Peptidilprolil Isomerase/imunologia , Proteínas Proto-Oncogênicas c-crk/imunologiaRESUMO
The immunogenicity and protective efficacy of recombinant SurA (rSurA) and rDnaK from Brucella spp. were evaluated in BALB/c mice. Immunization with rSurA in adjuvant induced a vigorous immunoglobulin G (IgG) response, with higher IgG2a than IgG1 titers. In addition, after in vitro stimulation with rSurA, spleen cells from rSurA-immunized mice produced interleukin-2 (IL-2), interferon (IFN)-gamma, IL-4 and IL-5. Immunization with rDnaK plus adjuvant induced a strong humoral response resulting in similar anti-rDnaK IgG titers than immunization with rDnaK alone. IgG2a titers predominated over IgG1 in mice injected with rDnaK alone or rDnaK plus adjuvant. Spleen cells from mice immunized with rDnaK plus adjuvant secreted IFN-gamma and IL-2 upon stimulation with rDnaK and induced a specific cytotoxic response. On the contrary, mice immunized with rDnaK alone did not exhibit a specific T helper or cytotoxic response in vitro. Mice given rSurA or rDnaK with adjuvant exhibited a significant degree of protection whereas immunization with rDnaK alone induced a low but still statistically significant level of protection against B. abortus infection. All studied vaccines were less protected than mice immunized with H38 or B. abortus strain 19 control vaccines. Altogether these results suggest that rSurA or rDnaK induce partial protection against B. abortus infection and could be useful candidates for the development of subunit vaccines against brucellosis.
Assuntos
Adenosina Trifosfatases/imunologia , Vacina contra Brucelose/imunologia , Brucella abortus/imunologia , Brucelose/imunologia , Brucelose/prevenção & controle , Proteínas de Transporte/imunologia , Peptidilprolil Isomerase/imunologia , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Animais , Formação de Anticorpos/imunologia , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Peptidilprolil Isomerase/genética , Peptidilprolil Isomerase/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Linfócitos T Citotóxicos/imunologia , Células Th1/imunologia , Células Th2/imunologiaRESUMO
We report the development of a novel protein-based nasal vaccine against Streptococcus pneumoniae, in which three pneumococcal proteins were displayed on the surface of a non-recombinant, killed Lactococcus lactis-derived delivery system, called Gram-positive Enhancer Matrix (GEM). The GEM particles induced the production of the proinflammatory cytokine tumour necrosis factor-alpha (TNF-alpha) by macrophages as well as the maturation of dendritic cells. The pneumococcal proteins IgA1 protease (IgA1p), putative proteinase maturation protein A (PpmA) and streptococcal lipoprotein A (SlrA) were anchored in trans to the surface of the GEM particles after recombinant production of the antigens in L. lactis as hybrids with a lactococcal cell wall binding domain, named Protein Anchor domain (PA). Intranasal immunisation with the SlrA-IgA1p or trivalent vaccine combinations without additional adjuvants showed significant protection against fatal pneumococcal pneumonia in mice. The GEM-based trivalent vaccine is a potential pneumococcal vaccine candidate that is expected to be easy to administer, safe and affordable to produce.
Assuntos
Lactococcus lactis/imunologia , Vacinas Pneumocócicas/imunologia , Streptococcus pneumoniae/imunologia , Administração Intranasal , Animais , Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Movimento Celular/imunologia , Células Dendríticas/imunologia , Feminino , Imunoglobulina G/imunologia , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Chaperonas Moleculares/imunologia , Peptidilprolil Isomerase/imunologia , Infecções Pneumocócicas/imunologia , Infecções Pneumocócicas/prevenção & controle , Vacinas Pneumocócicas/administração & dosagem , Vacinas Pneumocócicas/genética , Serina Endopeptidases/imunologia , Serina Endopeptidases/metabolismo , Streptococcus pneumoniae/genéticaRESUMO
Heat shock proteins (hsps) and cyclophilins (CypA) are intracellular chaperone molecules that facilitate protein folding and assembly. These proteins are selectively expressed in cells following exposure to a range of stress stimuli, including viral infection. Hsp species are highly immunogenic, eliciting humoral, cytotoxic T lymphocyte (CTL), and natural killer (NK) cell responses against viruses, tumours, and infectious diseases. This review discusses the roles of stress proteins in immunity and viral life cycles, vis-à-vis the development of Hsp-based therapeutic strategies against human immunodeficiency virus type-1 (HIV-1) infection. Cumulative findings are cited implicating the requirement of CypA in HIV-1 replication and formation of infectious virions. Studies by our group show the upregulated expression of hsp27 and hsp70 during single-cycle HIV infections. These species redistribute to the cell surface following HIV-infection and heat stress, serving as targets for NK and antibody-dependent cellular cytotoxicity. Co-immunoprecipitation and Western blot studies show that hsp27, hsp70, and hsp78 complex with HIV-1 viral proteins intracellularly. Hsp70, hsp56, and CypA are assembled into HIV-1 virions. The ability of hsps to interact with HIV-1 viral proteins, combined with their inherent adjuvant and immunogenic properties, indicates that hsps may serve as vehicles for antigen delivery and the design of vaccines against acquired immunodeficiency syndrome.
Assuntos
Infecções por HIV/prevenção & controle , HIV-1/imunologia , Proteínas de Choque Térmico/imunologia , Peptidilprolil Isomerase/imunologia , Humanos , VacinasRESUMO
TNF-alpha and LT-alpha are thought to be involved in the immunopathology of CNS demyelinating diseases. Both cytokines induce cellular effects through 55-kDa type-1 receptors (R1) and 75-kDa type-2 receptors (R2). To date, no study has specifically identified the various cell populations that express TNF receptors (TNFR) in the inflammatory and demyelinating mouse model, EAE. Phenotyping the TNFR positive cells is important in determining when and where the ligands may be acting and playing a role in disease pathology. We observed an upregulation of TNF R1 and R2 mRNA in high endothelial venules (HEVs) in the lymph node and CNS before the onset of EAE (preclinical phase). This upregulation of TNFR expression in HEVs was followed by a rapid increase in leukocytes within the CNS after the onset of clinical disease. The temporal kinetics of these data suggest that HEVs become activated early, probably through the release of pro-inflammatory cytokines originating from circulating leukocytes. An increase in TNFR on HEVs would make these cells more susceptible to TNF-induced changes, such as increasing cellular adhesion molecules, thereby further facilitating the trafficking of leukocytes into the CNS parenchyma.