RESUMO
Salmonella enterica is a major human pathogen that causes invasive non-typhoidal Salmonellosis (iNTS), resulting in significant morbidity and mortality. Although a number of pre-clinical and clinical studies have reported on the feasibility of developing a safe and effective vaccine against iNTS, there have been no licensed Salmonella vaccines available to protect against NTS strains. Vaccine formulations of highest priority for NTS are live attenuated vaccines, which can elicit effective induction of intestinal mucosal and intracellular bacteria-specific cell mediated immune responses. Since glucose is crucial for intracellular survival and replication in host cells, we constructed strains with mutations in components of the glucose uptake system, called the phosphotransferase system (PTS), and compared the relative virulence and immune responses in mice. In this study, we found that the strain with mutations in both ptsI and crr (KST0556) was the most attenuated strain among the tested strains, and proved to be highly effective in inducing a mucosal immune response that can protect against NTS infections in mice. Thus, we suggest here that KST0556 (ΔptsIΔcrr) is a potential live vaccine candidate for NTS, and may also be a candidate for a live delivery vector for heterologous antigens. Moreover, since PTS is a well-conserved glucose transporter system in both Gramnegative and Gram-positive bacteria, the ptsI and crr genes may be potential targets for creating live bacterial vectors or vaccine strains.
Assuntos
Proteínas de Bactérias/imunologia , Desoxirribonucleases de Sítio Específico do Tipo II/imunologia , Sistema Fosfotransferase de Açúcar do Fosfoenolpiruvato/imunologia , Infecções por Salmonella/prevenção & controle , Vacinas contra Salmonella/imunologia , Salmonella typhimurium/enzimologia , Vacinas Atenuadas/imunologia , Animais , Proteínas de Bactérias/administração & dosagem , Proteínas de Bactérias/genética , Desoxirribonucleases de Sítio Específico do Tipo II/administração & dosagem , Desoxirribonucleases de Sítio Específico do Tipo II/genética , Deleção de Genes , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Sistema Fosfotransferase de Açúcar do Fosfoenolpiruvato/administração & dosagem , Sistema Fosfotransferase de Açúcar do Fosfoenolpiruvato/genética , Infecções por Salmonella/imunologia , Infecções por Salmonella/microbiologia , Vacinas contra Salmonella/administração & dosagem , Vacinas contra Salmonella/genética , Salmonella typhimurium/genética , Salmonella typhimurium/imunologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genéticaRESUMO
BET inhibitors (BETi) target bromodomain-containing proteins and are currently being evaluated as anti-cancer agents. We find that maximal therapeutic effects of BETi in a Myc-driven B cell lymphoma model required an intact host immune system. Genome-wide analysis of the BETi-induced transcriptional response identified the immune checkpoint ligand Cd274 (Pd-l1) as a Myc-independent, BETi target-gene. BETi directly repressed constitutively expressed and interferon-gamma (IFN-γ) induced CD274 expression across different human and mouse tumor cell lines and primary patient samples. Mechanistically, BETi decreased Brd4 occupancy at the Cd274 locus without any change in Myc occupancy, resulting in transcriptional pausing and rapid loss of Cd274 mRNA production. Finally, targeted inhibition of the PD-1/PD-L1 axis by combining anti-PD-1 antibodies and the BETi JQ1 caused synergistic responses in mice bearing Myc-driven lymphomas. Our data uncover an interaction between BETi and the PD-1/PD-L1 immune-checkpoint and provide mechanistic insight into the transcriptional regulation of CD274.
Assuntos
Desoxirribonucleases de Sítio Específico do Tipo II/imunologia , Sistema Imunitário/imunologia , Proteínas Nucleares/imunologia , Receptor de Morte Celular Programada 1/imunologia , Fatores de Transcrição/imunologia , Animais , Antígeno B7-H1/imunologia , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Interferon gama/imunologia , Ligantes , Linfoma de Células B/imunologia , Camundongos , Proteínas Proto-Oncogênicas c-myc/imunologia , RNA Mensageiro/imunologia , Transcrição Gênica/imunologiaAssuntos
Desoxirribonucleases de Sítio Específico do Tipo II/isolamento & purificação , Anticorpos Monoclonais , Chlorella/virologia , Cromatografia de Afinidade/métodos , Desoxirribonucleases de Sítio Específico do Tipo II/imunologia , Guanidina/farmacologia , Imunoadsorventes/química , Phycodnaviridae/enzimologia , Sefarose/análogos & derivados , Sefarose/química , Ureia/farmacologiaRESUMO
The BamHI restriction-modification system contains a third gene, bamHIC, which positively regulates bamHIR. Similar small genes from other systems were tested in vivo for their ability to cross-complement. C.BamHI protein was identified, purified, and used to raise polyclonal antibodies. Attempts to detect other C proteins in cell extracts by cross-reactivity with C.BamHI antibodies proved unsuccessful.
Assuntos
Proteínas de Bactérias/genética , Enzimas de Restrição-Modificação do DNA/genética , Desoxirribonuclease BamHI , Genes Bacterianos/genética , Genes Reguladores , Bacillus/genética , Bacillus/imunologia , Proteínas de Bactérias/imunologia , Western Blotting , Sistema Livre de Células , Reações Cruzadas , Enzimas de Restrição-Modificação do DNA/imunologia , Desoxirribonucleases de Sítio Específico do Tipo II/genética , Desoxirribonucleases de Sítio Específico do Tipo II/imunologia , Teste de Complementação Genética , Biossíntese de Proteínas , Proteus vulgaris/genética , Proteus vulgaris/imunologia , Especificidade da Espécie , Transcrição GênicaRESUMO
The cross-reactivity of the monoclonal anti-human placental DNA methyltransferase antibody M2B10 with DNA methyltransferases isolated from other species was investigated. This antibody immunoprecipitates DNA methyltransferases from mammalian cells, i.e., human placenta, mouse P815 cells, and rat liver cells. No cross-reactivity is observed with DNA methyltransferases from wheat germ and with bacterial DNA methyltransferases HpaII and EcoRI. The mammalian enzymes are characterized by polypeptides of molecular mass 150-190 kDa. Polypeptides smaller than 190 kDa are presumably generated by proteolysis of the native 190-kDa DNA methyltransferase. Trypsin digestion of the 190-kDa polypeptide isolated from mouse cells results in progressive appearance of DNA methyltransferase polypeptides of 150-190, 110, 100, and 52-60 kDa.
