ABSTRACT
Deinococcus radiodurans (DR) is an extremophile that is well known for its resistance to radiation, oxidants and desiccation. The gene dr1790 of D. radiodurans was predicted to encode a yellow-related protein. The primary objective of the present study was to characterize the biological function of the DR1790 protein, which is a member of the ancient yellow/major royal jelly (MRJ) protein family, in prokaryotes. Fluorescence labeling demonstrated that the yellow-related protein encoded by dr1790 is a membrane protein. The deletion of the dr1790 gene decreased the cell growth rate and sensitivity to hydrogen peroxide and radiation and increased the membrane permeability of D. radiodurans. Transcript profiling by microarray and RT-PCR analyses of the dr1790 deletion mutant suggested that some genes that are involved in protein secretion and transport were strongly suppressed, while other genes that are involved in protein quality control, such as chaperones and proteases, were induced. In addition, the expression of genes with predicted functions that are involved in antioxidant systems, electron transport, and energy metabolism was significantly altered through the disruption of dr1790. Moreover, the results of proteomic analyses using 2-DE and MS also demonstrated that DR1790 contributed to D. radiodurans survival. Taken together, these results indicate that the DR1790 protein from the ancient yellow protein family plays a pleiotropic role in the survival of prokaryotic cells and contributes to the extraordinary resistance of D. radiodurans against oxidative and radiation stresses.
Subject(s)
Deinococcus/genetics , Genes, Bacterial , Genetic Pleiotropy , Mutagenesis, Insertional , Bacterial Proteins/genetics , Cell Membrane/physiology , Deinococcus/drug effects , Deinococcus/growth & development , Deinococcus/radiation effects , Gene Deletion , Gene Expression Profiling , Genetic Complementation Test , Hydrogen Peroxide/toxicity , Membrane Proteins/genetics , Microarray Analysis , Microbial Viability/drug effects , Microbial Viability/radiation effects , Permeability , Radiation, Ionizing , Real-Time Polymerase Chain ReactionABSTRACT
Deinococcus radiodurans (DR) is an extremophile that is well known for its resistance to radiation, oxidants and desiccation. The gene dr1790 of D. radiodurans was predicted to encode a yellow-related protein. The primary objective of the present study was to characterize the biological function of the DR1790 protein, which is a member of the ancient yellow/major royal jelly (MRJ) protein family, in prokaryotes. Fluorescence labeling demonstrated that the yellow-related protein encoded by dr1790 is a membrane protein. The deletion of the dr1790 gene decreased the cell growth rate and sensitivity to hydrogen peroxide and radiation and increased the membrane permeability of D. radiodurans. Transcript profiling by microarray and RT-PCR analyses of the dr1790 deletion mutant suggested that some genes that are involved in protein secretion and transport were strongly suppressed, while other genes that are involved in protein quality control, such as chaperones and proteases, were induced. In addition, the expression of genes with predicted functions that are involved in antioxidant systems, electron transport, and energy metabolism was significantly altered through the disruption of dr1790. Moreover, the results of proteomic analyses using 2-DE and MS also demonstrated that DR1790 contributed to D. radiodurans survival. Taken together, these results indicate that the DR1790 protein from the ancient yellow protein family plays a pleiotropic role in the survival of prokaryotic cells and contributes to the extraordinary resistance of D. radiodurans against oxidative and radiation stresses.
Subject(s)
Deinococcus/genetics , Genes, Bacterial , Genetic Pleiotropy , Mutagenesis, Insertional , Bacterial Proteins/genetics , Cell Membrane/physiology , Deinococcus/drug effects , Deinococcus/growth & development , Deinococcus/radiation effects , Gene Deletion , Gene Expression Profiling , Genetic Complementation Test , Hydrogen Peroxide/toxicity , Microarray Analysis , Membrane Proteins/genetics , Microbial Viability/drug effects , Microbial Viability/radiation effects , Permeability , Radiation, Ionizing , Real-Time Polymerase Chain ReactionABSTRACT
Deinococcus radiodurans (DR) is an extremophile that is well known for its resistance to radiation, oxidants and desiccation. The gene dr1790 of D. radiodurans was predicted to encode a yellow-related protein. The primary objective of the present study was to characterize the biological function of the DR1790 protein, which is a member of the ancient yellow/major royal jelly (MRJ) protein family, in prokaryotes. Fluorescence labeling demonstrated that the yellow-related protein encoded by dr1790 is a membrane protein. The deletion of the dr1790 gene decreased the cell growth rate and sensitivity to hydrogen peroxide and radiation and increased the membrane permeability of D. radiodurans. Transcript profiling by microarray and RT-PCR analyses of the dr1790 deletion mutant suggested that some genes that are involved in protein secretion and transport were strongly suppressed, while other genes that are involved in protein quality control, such as chaperones and proteases, were induced. In addition, the expression of genes with predicted functions that are involved in antioxidant systems, electron transport, and energy metabolism was significantly altered through the disruption of dr1790. Moreover, the results of proteomic analyses using 2-DE and MS also demonstrated that DR1790 contributed to D. radiodurans survival. Taken together, these results indicate that the DR1790 protein from the ancient yellow protein family plays a pleiotropic role in the survival of prokaryotic cells and contributes to the extraordinary resistance of D. radiodurans against oxidative and radiation stresses.(AU)
Subject(s)
Deinococcus/genetics , Genes, Bacterial , Genetic Pleiotropy , Mutagenesis, Insertional , Bacterial Proteins/genetics , Cell Membrane/physiology , Deinococcus , Deinococcus/growth & development , Deinococcus/radiation effects , Gene Deletion , Gene Expression Profiling , Genetic Complementation Test , Hydrogen Peroxide/toxicity , Membrane Proteins/genetics , Microarray Analysis , Microbial Viability , Microbial Viability/radiation effects , Permeability , Radiation, Ionizing , Real-Time Polymerase Chain ReactionABSTRACT
OBJECTIVE: Metallothionein 3 (MT-3) has been shown to protect against apoptotic neuronal death in the brains of patients with Alzheimer's disease. Zinc is a potent inhibitor of caspase-3 and its deficiency was found to promote apoptosis. Here, we measured the zinc and copper content in the brains of senescence-accelerated mouse/PRONE8 (SAMP8) and sought to investigate the effect of MT-3 on the apoptosis of neurons in the hippocampal CA1 region of these mice. METHOD: The zinc and copper content in the brain samples of SAMP8 and normal control SAMR1 mice were determined using an atomic absorption spectrophotometer. The mice were administered intraperitoneally for four weeks with MT-3 or MT1 and thereafter apoptosis was measured using the TUNEL method and the expression of anti-apoptotic protein Bcl-2 and proapoptotic protein Bax was examined by immunohistochemistry. RESULTS: Compared with that in SMAR1 mice, the content of zinc in the brains of SAMP8 mice was significantly reduced (P<0.05). Moreover, significant levels of apoptosis of neurons were observed in the hippocampus of SAMP8 mice, which, compared with those in SMAR1 mice, also showed significantly lower levels of Bcl-2 and higher levels of Bax (P<0.05). MT-3 increased zinc concentration in the hippocampus of SAMP8 mice and also significantly decreased apoptosis in these neurons dose-dependently and increased the levels of Bcl-2 and decreased the levels of Bax. CONCLUSION: MT-3 could attenuate apoptotic neuron death in the hippocampus of SAMP8, suggesting that the protein may lessen the development of neurodegeneration.
Subject(s)
Apoptosis/drug effects , Growth Inhibitors/pharmacology , Hippocampus/drug effects , Nerve Tissue Proteins/pharmacology , Neurons/drug effects , Aging , Animals , Brain Chemistry , Caspase 3/deficiency , Caspase Inhibitors , Copper/analysis , Hippocampus/metabolism , Hippocampus/pathology , Male , Metallothionein/pharmacology , Metallothionein 3 , Mice , Neurons/metabolism , Neurons/pathology , Proto-Oncogene Proteins c-bcl-2/analysis , Species Specificity , Zinc/analysis , Zinc/deficiency , bcl-2-Associated X Protein/analysisABSTRACT
OBJECTIVE: Metallothionein 3 (MT-3) has been shown to protect against apoptotic neuronal death in the brains of patients with Alzheimer's disease. Zinc is a potent inhibitor of caspase-3 and its deficiency was found to promote apoptosis. Here, we measured the zinc and copper content in the brains of senescence-accelerated mouse/PRONE8 (SAMP8) and sought to investigate the effect of MT-3 on the apoptosis of neurons in the hippocampal CA1 region of these mice. METHOD: The zinc and copper content in the brain samples of SAMP8 and normal control SAMR1 mice were determined using an atomic absorption spectrophotometer. The mice were administered intraperitoneally for four weeks with MT-3 or MT1 and thereafter apoptosis was measured using the TUNEL method and the expression of anti-apoptotic protein Bcl-2 and proapoptotic protein Bax was examined by immunohistochemistry. RESULTS: Compared with that in SMAR1 mice, the content of zinc in the brains of SAMP8 mice was significantly reduced (P<0.05). Moreover, significant levels of apoptosis of neurons were observed in the hippocampus of SAMP8 mice, which, compared with those in SMAR1 mice, also showed significantly lower levels of Bcl-2 and higher levels of Bax (P<0.05). MT-3 increased zinc concentration in the hippocampus of SAMP8 mice and also significantly decreased apoptosis in these neurons dose-dependently and increased the levels of Bcl-2 and decreased the levels of Bax. CONCLUSION: MT-3 could attenuate apoptotic neuron death in the hippocampus of SAMP8, suggesting that the protein may lessen the development of neurodegeneration.
OBJETIVO: Metalotioneína 3 (MT-3) tem mostrado proteção contra a apoptose neuronal em cérebros de pacientes com doença de Alzheimer. Zinco é um potente inibidor da caspase-3, e sua deficiência pode promover a apoptose. No presente trabalho, foram dosados os níveis de zinco e cobre nos cérebros de camundongos PRONE8 com envelhecimento acelerado (SAMP8), visando investigar o efeito da MT-3 na apoptse dos neurônios da região hipocampal CA1 destes camundongos. MÉTODO: Os níveis de zinco e cobre em amostras cerebrais de camundongos SAMP8 e de controles normais SAMR1 foram determinados por absorção atômica em espectrofotometria. Foram administradas MT-3 ou MT-1 intraperitoneais durante quatro semanas, sendo em seguida avaliada a apoptose pelo método TUNEL , enquanto a expressão da proteína anti-apoptótica Bcl-2 e a proteína pró-apoptótica Bax foram avaliadas por imunohistoquímica. RESULTADOS: Em comparação aos camundongos SMAR1, o nível de zinco nas amostras cerebrais dos camundongos SAMP8 estava significativamente diminuído (P<0.05). Além disto, níveis significativos de apoptose foram observados no hipocampo dos camundongos SAMP8, o que, em comparação com os níveis em camundongos SMAR1, também mostrava níveis significativamente mais baixos de Bcl-2 e níveis mais altos de Bax (P<0.05). MT-3 aumentou a concentração de zinco no hipocampo dos camundongos SAMP8, além de diminuir significativamente a apoptose destes neurônios, de uma forma dose-dependente, ao mesmo tempo que aumentou níveis de Bcl-2 e diminuiu níveis de Bax. CONCLUSÃO: MT-3 pode atenuar a morte neuronal apoptótica no hipocampo de SAMP8, o que sugere que esta proteína possa diminuir a neurodegeneração.