The Experts below are selected from a list of 249 Experts worldwide ranked by ideXlab platform
Wulf Paschen - One of the best experts on this subject based on the ideXlab platform.
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Small Ubiquitin-Like Modifier 3–Modified Proteome Regulated by Brain Ischemia in Novel Small Ubiquitin-Like Modifier Transgenic Mice: Putative Protective Proteins/Pathways
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, Pei Miao, Thompson Jw, Moseley Ma, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose— Small ubiquitin-like modifier (SUMO) conjugation is a post-translational modification associated with many human diseases. Characterization of the SUMO-modified proteome is pivotal to define the mechanistic link between SUMO conjugation and such diseases. This is particularly evident for SUMO2/3 conjugation, which is massively activated after brain ischemia/stroke, and is believed to be a protective response. The purpose of this study was to perform a comprehensive analysis of the SUMO3-modified proteome regulated by brain ischemia using a novel SUMO transgenic mouse. Methods— To enable SUMO proteomics analysis in vivo, we generated transgenic mice conditionally expressing tagged SUMO1-3 paralogues. Transgenic mice were subjected to 10 minutes forebrain ischemia and 1 hour of reperfusion. SUMO3-Conjugated Proteins were enriched by anti-FLAG affinity purification and analyzed by liquid chromatography–tandem mass spectrometry. Results— Characterization of SUMO transgenic mice demonstrated that all 3 tagged SUMO paralogues were functionally active, and expression of exogenous SUMOs did not modify the endogenous SUMOylation machinery. Proteomics analysis identified 112 putative SUMO3 substrates of which 91 candidates were more abundant in the ischemia group than the sham group. Data analysis revealed processes/pathways with putative neuroprotective functions, including glucocorticoid receptor signaling, RNA processing, and SUMOylation-dependent ubiquitin conjugation. Conclusions— The identified Proteins/pathways modulated by SUMOylation could be the key to understand the mechanisms linking SUMOylation to neuroprotection, and thus provide new promising targets for therapeutic interventions. The new transgenic mouse will be an invaluable platform for analyzing the SUMO-modified proteome in models of human disorders and thereby help to mechanistically link SUMOylation to the pathological processes.
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abstract w p206 analysis of sumo3 modified proteome in post ischemic mouse brain
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, William M Thompson, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose: Small ubiquitin-like modifier (SUMO) conjugation modulates many key cellular processes. Transient cerebral ischemia dramatically activates SUMO2/3 conjugation, and this is believed to be a protective stress response. It is, therefore, of tremendous clinical interest to characterize the SUMO-modified proteome regulated by transient ischemia. We generated a novel SUMO transgenic mouse and performed the first SUMO proteomics study using post-ischemic brain samples. Methods: CAG-loxP-STOP-loxP-SUMO (CAG-SUMO) mice were generated in which His-SUMO1, HA-SUMO2, and FLAG-SUMO3 were expressed from a single multicistronic transgene in a Cre-dependent manner. CAG-SUMO mice were mated with Emx1 Cre/Cre mice to generate double transgenic CAG-SUMO/Emx1-Cre mice as experimental mice and Emx1 Cre/+ mice as control mice. Double transgenic mice were subjected to 10 min global cerebral ischemia followed by 1 h reperfusion or sham operation. FLAG-SUMO3-Conjugated Proteins were enriched from cortical tissues and analyzed. Results: Characterization of double transgenic mice demonstrated that exogenous expressed tagged SUMO paralogues were functionally intact and did not perturb the endogenous SUMOylation machinery in the brain. FLAG pulldown of cortical samples from sham and ischemia mice followed by GeLC-MS/MS analysis identified 91 candidates whose SUMOylation states were up-regulated in ischemic samples. Data analysis revealed several potentially important processes in which SUMO3 conjugation may play a key role during ischemia/reperfusion, including the cross-talk between SUMOylation and ubiquitination, glucocorticoid receptor signaling, and modulation of posttranscriptional mRNA processing. Conclusions: SUMO proteomic analysis identified important processes and pathways modulated by SUMOylation in the post-ischemic brain that warrant future investigations, since they could be the key to understand the overall impact of SUMOylation on the fate and functions of post-ischemic neurons. The conditional SUMO transgenic mouse will be an invaluable tool for in-depth in vivo analysis of the SUMO-modified proteome in various pathological states.
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transient ischemia induces massive nuclear accumulation of sumo2 3 Conjugated Proteins in spinal cord neurons
Spinal Cord, 2013Co-Authors: Wulf Paschen, Huaxin Sheng, Z Wang, R Wang, S P Sheng, Wei YangAbstract:Transient ischemia induces massive nuclear accumulation of SUMO2/3-Conjugated Proteins in spinal cord neurons
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moderate hypothermia induces marked increase in levels and nuclear accumulation of sumo2 3 Conjugated Proteins in neurons
Journal of Neurochemistry, 2012Co-Authors: Liangli Wang, Wei Yang, Burkhard G Mackensen, Wulf PaschenAbstract:Deep hypothermia protects the brain from ischemic damage and is therefore used during major cardiovascular surgeries requiring cardiopulmonary bypass and a period of circulatory arrest. Here, we demonstrated that small ubiquitin-like modifier (SUMO1-3) conjugation is markedly activated in the brain during deep to moderate hypothermia. Animals were subjected to normothermic (37°C) or deep to moderate (18°C, 24°C, 30°C) hypothermic cardiopulmonary bypass, and the effects of hypothermia on SUMO conjugation were evaluated by Western blot and immunohistochemistry. Exposure to moderate 30°C hypothermia was sufficient to markedly increase levels and nuclear accumulation of SUMO2/3-Conjugated Proteins in these cells. Deep hypothermia induced nuclear translocation of the SUMO-conjugating enzyme Ubc9, suggesting that the increase in nuclear levels of SUMO2/3-Conjugated Proteins observed in brains of hypothermic animals is an active process. Exposure of primary neuronal cultures to deep hypothermia induced only a moderate rise in levels of SUMO2/3-Conjugated Proteins. This suggests that neurons in vivo have a higher capacity than neurons in vitro to activate this endogenous potentially neuroprotective pathway upon exposure to hypothermia. Identifying Proteins that are SUMO2/3 Conjugated during hypothermia could help to design new strategies for preventive and therapeutic interventions to make neurons more resistant to a transient interruption of blood supply.
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Moderate hypothermia induces marked increase in levels and nuclear accumulation of SUMO2/3‐Conjugated Proteins in neurons
Journal of Neurochemistry, 2012Co-Authors: Liangli Wang, Wei Yang, Qing Ma, G. Burkhard Mackensen, Wulf PaschenAbstract:Deep hypothermia protects the brain from ischemic damage and is therefore used during major cardiovascular surgeries requiring cardiopulmonary bypass and a period of circulatory arrest. Here, we demonstrated that small ubiquitin-like modifier (SUMO1-3) conjugation is markedly activated in the brain during deep to moderate hypothermia. Animals were subjected to normothermic (37°C) or deep to moderate (18°C, 24°C, 30°C) hypothermic cardiopulmonary bypass, and the effects of hypothermia on SUMO conjugation were evaluated by Western blot and immunohistochemistry. Exposure to moderate 30°C hypothermia was sufficient to markedly increase levels and nuclear accumulation of SUMO2/3-Conjugated Proteins in these cells. Deep hypothermia induced nuclear translocation of the SUMO-conjugating enzyme Ubc9, suggesting that the increase in nuclear levels of SUMO2/3-Conjugated Proteins observed in brains of hypothermic animals is an active process. Exposure of primary neuronal cultures to deep hypothermia induced only a moderate rise in levels of SUMO2/3-Conjugated Proteins. This suggests that neurons in vivo have a higher capacity than neurons in vitro to activate this endogenous potentially neuroprotective pathway upon exposure to hypothermia. Identifying Proteins that are SUMO2/3 Conjugated during hypothermia could help to design new strategies for preventive and therapeutic interventions to make neurons more resistant to a transient interruption of blood supply.
Wei Yang - One of the best experts on this subject based on the ideXlab platform.
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Small Ubiquitin-Like Modifier 3–Modified Proteome Regulated by Brain Ischemia in Novel Small Ubiquitin-Like Modifier Transgenic Mice: Putative Protective Proteins/Pathways
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, Pei Miao, Thompson Jw, Moseley Ma, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose— Small ubiquitin-like modifier (SUMO) conjugation is a post-translational modification associated with many human diseases. Characterization of the SUMO-modified proteome is pivotal to define the mechanistic link between SUMO conjugation and such diseases. This is particularly evident for SUMO2/3 conjugation, which is massively activated after brain ischemia/stroke, and is believed to be a protective response. The purpose of this study was to perform a comprehensive analysis of the SUMO3-modified proteome regulated by brain ischemia using a novel SUMO transgenic mouse. Methods— To enable SUMO proteomics analysis in vivo, we generated transgenic mice conditionally expressing tagged SUMO1-3 paralogues. Transgenic mice were subjected to 10 minutes forebrain ischemia and 1 hour of reperfusion. SUMO3-Conjugated Proteins were enriched by anti-FLAG affinity purification and analyzed by liquid chromatography–tandem mass spectrometry. Results— Characterization of SUMO transgenic mice demonstrated that all 3 tagged SUMO paralogues were functionally active, and expression of exogenous SUMOs did not modify the endogenous SUMOylation machinery. Proteomics analysis identified 112 putative SUMO3 substrates of which 91 candidates were more abundant in the ischemia group than the sham group. Data analysis revealed processes/pathways with putative neuroprotective functions, including glucocorticoid receptor signaling, RNA processing, and SUMOylation-dependent ubiquitin conjugation. Conclusions— The identified Proteins/pathways modulated by SUMOylation could be the key to understand the mechanisms linking SUMOylation to neuroprotection, and thus provide new promising targets for therapeutic interventions. The new transgenic mouse will be an invaluable platform for analyzing the SUMO-modified proteome in models of human disorders and thereby help to mechanistically link SUMOylation to the pathological processes.
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abstract w p206 analysis of sumo3 modified proteome in post ischemic mouse brain
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, William M Thompson, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose: Small ubiquitin-like modifier (SUMO) conjugation modulates many key cellular processes. Transient cerebral ischemia dramatically activates SUMO2/3 conjugation, and this is believed to be a protective stress response. It is, therefore, of tremendous clinical interest to characterize the SUMO-modified proteome regulated by transient ischemia. We generated a novel SUMO transgenic mouse and performed the first SUMO proteomics study using post-ischemic brain samples. Methods: CAG-loxP-STOP-loxP-SUMO (CAG-SUMO) mice were generated in which His-SUMO1, HA-SUMO2, and FLAG-SUMO3 were expressed from a single multicistronic transgene in a Cre-dependent manner. CAG-SUMO mice were mated with Emx1 Cre/Cre mice to generate double transgenic CAG-SUMO/Emx1-Cre mice as experimental mice and Emx1 Cre/+ mice as control mice. Double transgenic mice were subjected to 10 min global cerebral ischemia followed by 1 h reperfusion or sham operation. FLAG-SUMO3-Conjugated Proteins were enriched from cortical tissues and analyzed. Results: Characterization of double transgenic mice demonstrated that exogenous expressed tagged SUMO paralogues were functionally intact and did not perturb the endogenous SUMOylation machinery in the brain. FLAG pulldown of cortical samples from sham and ischemia mice followed by GeLC-MS/MS analysis identified 91 candidates whose SUMOylation states were up-regulated in ischemic samples. Data analysis revealed several potentially important processes in which SUMO3 conjugation may play a key role during ischemia/reperfusion, including the cross-talk between SUMOylation and ubiquitination, glucocorticoid receptor signaling, and modulation of posttranscriptional mRNA processing. Conclusions: SUMO proteomic analysis identified important processes and pathways modulated by SUMOylation in the post-ischemic brain that warrant future investigations, since they could be the key to understand the overall impact of SUMOylation on the fate and functions of post-ischemic neurons. The conditional SUMO transgenic mouse will be an invaluable tool for in-depth in vivo analysis of the SUMO-modified proteome in various pathological states.
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transient ischemia induces massive nuclear accumulation of sumo2 3 Conjugated Proteins in spinal cord neurons
Spinal Cord, 2013Co-Authors: Wulf Paschen, Huaxin Sheng, Z Wang, R Wang, S P Sheng, Wei YangAbstract:Transient ischemia induces massive nuclear accumulation of SUMO2/3-Conjugated Proteins in spinal cord neurons
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moderate hypothermia induces marked increase in levels and nuclear accumulation of sumo2 3 Conjugated Proteins in neurons
Journal of Neurochemistry, 2012Co-Authors: Liangli Wang, Wei Yang, Burkhard G Mackensen, Wulf PaschenAbstract:Deep hypothermia protects the brain from ischemic damage and is therefore used during major cardiovascular surgeries requiring cardiopulmonary bypass and a period of circulatory arrest. Here, we demonstrated that small ubiquitin-like modifier (SUMO1-3) conjugation is markedly activated in the brain during deep to moderate hypothermia. Animals were subjected to normothermic (37°C) or deep to moderate (18°C, 24°C, 30°C) hypothermic cardiopulmonary bypass, and the effects of hypothermia on SUMO conjugation were evaluated by Western blot and immunohistochemistry. Exposure to moderate 30°C hypothermia was sufficient to markedly increase levels and nuclear accumulation of SUMO2/3-Conjugated Proteins in these cells. Deep hypothermia induced nuclear translocation of the SUMO-conjugating enzyme Ubc9, suggesting that the increase in nuclear levels of SUMO2/3-Conjugated Proteins observed in brains of hypothermic animals is an active process. Exposure of primary neuronal cultures to deep hypothermia induced only a moderate rise in levels of SUMO2/3-Conjugated Proteins. This suggests that neurons in vivo have a higher capacity than neurons in vitro to activate this endogenous potentially neuroprotective pathway upon exposure to hypothermia. Identifying Proteins that are SUMO2/3 Conjugated during hypothermia could help to design new strategies for preventive and therapeutic interventions to make neurons more resistant to a transient interruption of blood supply.
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Moderate hypothermia induces marked increase in levels and nuclear accumulation of SUMO2/3‐Conjugated Proteins in neurons
Journal of Neurochemistry, 2012Co-Authors: Liangli Wang, Wei Yang, Qing Ma, G. Burkhard Mackensen, Wulf PaschenAbstract:Deep hypothermia protects the brain from ischemic damage and is therefore used during major cardiovascular surgeries requiring cardiopulmonary bypass and a period of circulatory arrest. Here, we demonstrated that small ubiquitin-like modifier (SUMO1-3) conjugation is markedly activated in the brain during deep to moderate hypothermia. Animals were subjected to normothermic (37°C) or deep to moderate (18°C, 24°C, 30°C) hypothermic cardiopulmonary bypass, and the effects of hypothermia on SUMO conjugation were evaluated by Western blot and immunohistochemistry. Exposure to moderate 30°C hypothermia was sufficient to markedly increase levels and nuclear accumulation of SUMO2/3-Conjugated Proteins in these cells. Deep hypothermia induced nuclear translocation of the SUMO-conjugating enzyme Ubc9, suggesting that the increase in nuclear levels of SUMO2/3-Conjugated Proteins observed in brains of hypothermic animals is an active process. Exposure of primary neuronal cultures to deep hypothermia induced only a moderate rise in levels of SUMO2/3-Conjugated Proteins. This suggests that neurons in vivo have a higher capacity than neurons in vitro to activate this endogenous potentially neuroprotective pathway upon exposure to hypothermia. Identifying Proteins that are SUMO2/3 Conjugated during hypothermia could help to design new strategies for preventive and therapeutic interventions to make neurons more resistant to a transient interruption of blood supply.
Liangli Wang - One of the best experts on this subject based on the ideXlab platform.
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Small Ubiquitin-Like Modifier 3–Modified Proteome Regulated by Brain Ischemia in Novel Small Ubiquitin-Like Modifier Transgenic Mice: Putative Protective Proteins/Pathways
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, Pei Miao, Thompson Jw, Moseley Ma, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose— Small ubiquitin-like modifier (SUMO) conjugation is a post-translational modification associated with many human diseases. Characterization of the SUMO-modified proteome is pivotal to define the mechanistic link between SUMO conjugation and such diseases. This is particularly evident for SUMO2/3 conjugation, which is massively activated after brain ischemia/stroke, and is believed to be a protective response. The purpose of this study was to perform a comprehensive analysis of the SUMO3-modified proteome regulated by brain ischemia using a novel SUMO transgenic mouse. Methods— To enable SUMO proteomics analysis in vivo, we generated transgenic mice conditionally expressing tagged SUMO1-3 paralogues. Transgenic mice were subjected to 10 minutes forebrain ischemia and 1 hour of reperfusion. SUMO3-Conjugated Proteins were enriched by anti-FLAG affinity purification and analyzed by liquid chromatography–tandem mass spectrometry. Results— Characterization of SUMO transgenic mice demonstrated that all 3 tagged SUMO paralogues were functionally active, and expression of exogenous SUMOs did not modify the endogenous SUMOylation machinery. Proteomics analysis identified 112 putative SUMO3 substrates of which 91 candidates were more abundant in the ischemia group than the sham group. Data analysis revealed processes/pathways with putative neuroprotective functions, including glucocorticoid receptor signaling, RNA processing, and SUMOylation-dependent ubiquitin conjugation. Conclusions— The identified Proteins/pathways modulated by SUMOylation could be the key to understand the mechanisms linking SUMOylation to neuroprotection, and thus provide new promising targets for therapeutic interventions. The new transgenic mouse will be an invaluable platform for analyzing the SUMO-modified proteome in models of human disorders and thereby help to mechanistically link SUMOylation to the pathological processes.
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abstract w p206 analysis of sumo3 modified proteome in post ischemic mouse brain
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, William M Thompson, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose: Small ubiquitin-like modifier (SUMO) conjugation modulates many key cellular processes. Transient cerebral ischemia dramatically activates SUMO2/3 conjugation, and this is believed to be a protective stress response. It is, therefore, of tremendous clinical interest to characterize the SUMO-modified proteome regulated by transient ischemia. We generated a novel SUMO transgenic mouse and performed the first SUMO proteomics study using post-ischemic brain samples. Methods: CAG-loxP-STOP-loxP-SUMO (CAG-SUMO) mice were generated in which His-SUMO1, HA-SUMO2, and FLAG-SUMO3 were expressed from a single multicistronic transgene in a Cre-dependent manner. CAG-SUMO mice were mated with Emx1 Cre/Cre mice to generate double transgenic CAG-SUMO/Emx1-Cre mice as experimental mice and Emx1 Cre/+ mice as control mice. Double transgenic mice were subjected to 10 min global cerebral ischemia followed by 1 h reperfusion or sham operation. FLAG-SUMO3-Conjugated Proteins were enriched from cortical tissues and analyzed. Results: Characterization of double transgenic mice demonstrated that exogenous expressed tagged SUMO paralogues were functionally intact and did not perturb the endogenous SUMOylation machinery in the brain. FLAG pulldown of cortical samples from sham and ischemia mice followed by GeLC-MS/MS analysis identified 91 candidates whose SUMOylation states were up-regulated in ischemic samples. Data analysis revealed several potentially important processes in which SUMO3 conjugation may play a key role during ischemia/reperfusion, including the cross-talk between SUMOylation and ubiquitination, glucocorticoid receptor signaling, and modulation of posttranscriptional mRNA processing. Conclusions: SUMO proteomic analysis identified important processes and pathways modulated by SUMOylation in the post-ischemic brain that warrant future investigations, since they could be the key to understand the overall impact of SUMOylation on the fate and functions of post-ischemic neurons. The conditional SUMO transgenic mouse will be an invaluable tool for in-depth in vivo analysis of the SUMO-modified proteome in various pathological states.
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moderate hypothermia induces marked increase in levels and nuclear accumulation of sumo2 3 Conjugated Proteins in neurons
Journal of Neurochemistry, 2012Co-Authors: Liangli Wang, Wei Yang, Burkhard G Mackensen, Wulf PaschenAbstract:Deep hypothermia protects the brain from ischemic damage and is therefore used during major cardiovascular surgeries requiring cardiopulmonary bypass and a period of circulatory arrest. Here, we demonstrated that small ubiquitin-like modifier (SUMO1-3) conjugation is markedly activated in the brain during deep to moderate hypothermia. Animals were subjected to normothermic (37°C) or deep to moderate (18°C, 24°C, 30°C) hypothermic cardiopulmonary bypass, and the effects of hypothermia on SUMO conjugation were evaluated by Western blot and immunohistochemistry. Exposure to moderate 30°C hypothermia was sufficient to markedly increase levels and nuclear accumulation of SUMO2/3-Conjugated Proteins in these cells. Deep hypothermia induced nuclear translocation of the SUMO-conjugating enzyme Ubc9, suggesting that the increase in nuclear levels of SUMO2/3-Conjugated Proteins observed in brains of hypothermic animals is an active process. Exposure of primary neuronal cultures to deep hypothermia induced only a moderate rise in levels of SUMO2/3-Conjugated Proteins. This suggests that neurons in vivo have a higher capacity than neurons in vitro to activate this endogenous potentially neuroprotective pathway upon exposure to hypothermia. Identifying Proteins that are SUMO2/3 Conjugated during hypothermia could help to design new strategies for preventive and therapeutic interventions to make neurons more resistant to a transient interruption of blood supply.
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Moderate hypothermia induces marked increase in levels and nuclear accumulation of SUMO2/3‐Conjugated Proteins in neurons
Journal of Neurochemistry, 2012Co-Authors: Liangli Wang, Wei Yang, Qing Ma, G. Burkhard Mackensen, Wulf PaschenAbstract:Deep hypothermia protects the brain from ischemic damage and is therefore used during major cardiovascular surgeries requiring cardiopulmonary bypass and a period of circulatory arrest. Here, we demonstrated that small ubiquitin-like modifier (SUMO1-3) conjugation is markedly activated in the brain during deep to moderate hypothermia. Animals were subjected to normothermic (37°C) or deep to moderate (18°C, 24°C, 30°C) hypothermic cardiopulmonary bypass, and the effects of hypothermia on SUMO conjugation were evaluated by Western blot and immunohistochemistry. Exposure to moderate 30°C hypothermia was sufficient to markedly increase levels and nuclear accumulation of SUMO2/3-Conjugated Proteins in these cells. Deep hypothermia induced nuclear translocation of the SUMO-conjugating enzyme Ubc9, suggesting that the increase in nuclear levels of SUMO2/3-Conjugated Proteins observed in brains of hypothermic animals is an active process. Exposure of primary neuronal cultures to deep hypothermia induced only a moderate rise in levels of SUMO2/3-Conjugated Proteins. This suggests that neurons in vivo have a higher capacity than neurons in vitro to activate this endogenous potentially neuroprotective pathway upon exposure to hypothermia. Identifying Proteins that are SUMO2/3 Conjugated during hypothermia could help to design new strategies for preventive and therapeutic interventions to make neurons more resistant to a transient interruption of blood supply.
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analysis of oxygen glucose deprivation induced changes in sumo3 conjugation using silac based quantitative proteomics
Journal of Proteome Research, 2012Co-Authors: Wei Yang, Liangli Wang, Will J Thompson, Arthur M Moseley, Zhengfeng Wang, Matthew W. Foster, Huaxin Sheng, Wulf PaschenAbstract:Transient cerebral ischemia dramatically activates small ubiquitin-like modifier (SUMO2/3) conjugation. In cells exposed to 6 h of transient oxygen/glucose deprivation (OGD), a model of ischemia, SUMOylation increases profoundly between 0 and 30 min following re-oxygenation. To elucidate the effect of transient OGD on SUMO conjugation of target Proteins, we exposed neuroblastoma B35 cells expressing HA-SUMO3 to transient OGD and used stable isotope labeling with amino acids in cell culture (SILAC) to quantify OGD-induced changes in levels of specific SUMOylated Proteins. Lysates from control and OGD-treated cells were mixed equally, and HA-tagged Proteins were immunoprecipitated and analyzed by 1D-SDS-PAGE-LC-MS/MS. We identified 188 putative SUMO3-Conjugated Proteins, including numerous transcription factors and coregulators, and PIAS2 and PIAS4 SUMO ligases, of which 22 were increased or decreased >±2-fold. In addition to SUMO3, the levels of protein-Conjugated SUMO1 and SUMO2, as well as ubiquitin, were all increased. Importantly, protein ubiquitination induced by OGD was completely blocked by gene silencing of SUMO2/3. Collectively, these results suggest several mechanisms for OGD-modulated SUMOylation, point to a number of signaling pathways that may be targets of SUMO-based signaling and recovery from ischemic stress, as well as demonstrate a tightly controlled crosstalk between the SUMO and ubiquitin conjugation pathways.
Huaxin Sheng - One of the best experts on this subject based on the ideXlab platform.
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Small Ubiquitin-Like Modifier 3–Modified Proteome Regulated by Brain Ischemia in Novel Small Ubiquitin-Like Modifier Transgenic Mice: Putative Protective Proteins/Pathways
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, Pei Miao, Thompson Jw, Moseley Ma, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose— Small ubiquitin-like modifier (SUMO) conjugation is a post-translational modification associated with many human diseases. Characterization of the SUMO-modified proteome is pivotal to define the mechanistic link between SUMO conjugation and such diseases. This is particularly evident for SUMO2/3 conjugation, which is massively activated after brain ischemia/stroke, and is believed to be a protective response. The purpose of this study was to perform a comprehensive analysis of the SUMO3-modified proteome regulated by brain ischemia using a novel SUMO transgenic mouse. Methods— To enable SUMO proteomics analysis in vivo, we generated transgenic mice conditionally expressing tagged SUMO1-3 paralogues. Transgenic mice were subjected to 10 minutes forebrain ischemia and 1 hour of reperfusion. SUMO3-Conjugated Proteins were enriched by anti-FLAG affinity purification and analyzed by liquid chromatography–tandem mass spectrometry. Results— Characterization of SUMO transgenic mice demonstrated that all 3 tagged SUMO paralogues were functionally active, and expression of exogenous SUMOs did not modify the endogenous SUMOylation machinery. Proteomics analysis identified 112 putative SUMO3 substrates of which 91 candidates were more abundant in the ischemia group than the sham group. Data analysis revealed processes/pathways with putative neuroprotective functions, including glucocorticoid receptor signaling, RNA processing, and SUMOylation-dependent ubiquitin conjugation. Conclusions— The identified Proteins/pathways modulated by SUMOylation could be the key to understand the mechanisms linking SUMOylation to neuroprotection, and thus provide new promising targets for therapeutic interventions. The new transgenic mouse will be an invaluable platform for analyzing the SUMO-modified proteome in models of human disorders and thereby help to mechanistically link SUMOylation to the pathological processes.
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abstract w p206 analysis of sumo3 modified proteome in post ischemic mouse brain
Stroke, 2014Co-Authors: Wei Yang, Liangli Wang, William M Thompson, Shengli Zhao, Huaxin Sheng, Wulf PaschenAbstract:Background and Purpose: Small ubiquitin-like modifier (SUMO) conjugation modulates many key cellular processes. Transient cerebral ischemia dramatically activates SUMO2/3 conjugation, and this is believed to be a protective stress response. It is, therefore, of tremendous clinical interest to characterize the SUMO-modified proteome regulated by transient ischemia. We generated a novel SUMO transgenic mouse and performed the first SUMO proteomics study using post-ischemic brain samples. Methods: CAG-loxP-STOP-loxP-SUMO (CAG-SUMO) mice were generated in which His-SUMO1, HA-SUMO2, and FLAG-SUMO3 were expressed from a single multicistronic transgene in a Cre-dependent manner. CAG-SUMO mice were mated with Emx1 Cre/Cre mice to generate double transgenic CAG-SUMO/Emx1-Cre mice as experimental mice and Emx1 Cre/+ mice as control mice. Double transgenic mice were subjected to 10 min global cerebral ischemia followed by 1 h reperfusion or sham operation. FLAG-SUMO3-Conjugated Proteins were enriched from cortical tissues and analyzed. Results: Characterization of double transgenic mice demonstrated that exogenous expressed tagged SUMO paralogues were functionally intact and did not perturb the endogenous SUMOylation machinery in the brain. FLAG pulldown of cortical samples from sham and ischemia mice followed by GeLC-MS/MS analysis identified 91 candidates whose SUMOylation states were up-regulated in ischemic samples. Data analysis revealed several potentially important processes in which SUMO3 conjugation may play a key role during ischemia/reperfusion, including the cross-talk between SUMOylation and ubiquitination, glucocorticoid receptor signaling, and modulation of posttranscriptional mRNA processing. Conclusions: SUMO proteomic analysis identified important processes and pathways modulated by SUMOylation in the post-ischemic brain that warrant future investigations, since they could be the key to understand the overall impact of SUMOylation on the fate and functions of post-ischemic neurons. The conditional SUMO transgenic mouse will be an invaluable tool for in-depth in vivo analysis of the SUMO-modified proteome in various pathological states.
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transient ischemia induces massive nuclear accumulation of sumo2 3 Conjugated Proteins in spinal cord neurons
Spinal Cord, 2013Co-Authors: Wulf Paschen, Huaxin Sheng, Z Wang, R Wang, S P Sheng, Wei YangAbstract:Transient ischemia induces massive nuclear accumulation of SUMO2/3-Conjugated Proteins in spinal cord neurons
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Transient ischemia induces massive nuclear accumulation of SUMO2/3-Conjugated Proteins in spinal cord neurons
Spinal Cord, 2012Co-Authors: Z Wang, Wulf Paschen, Huaxin Sheng, R Wang, S P Sheng, Wei YangAbstract:Transient ischemia induces massive nuclear accumulation of SUMO2/3-Conjugated Proteins in spinal cord neurons
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analysis of oxygen glucose deprivation induced changes in sumo3 conjugation using silac based quantitative proteomics
Journal of Proteome Research, 2012Co-Authors: Wei Yang, Liangli Wang, Will J Thompson, Arthur M Moseley, Zhengfeng Wang, Matthew W. Foster, Huaxin Sheng, Wulf PaschenAbstract:Transient cerebral ischemia dramatically activates small ubiquitin-like modifier (SUMO2/3) conjugation. In cells exposed to 6 h of transient oxygen/glucose deprivation (OGD), a model of ischemia, SUMOylation increases profoundly between 0 and 30 min following re-oxygenation. To elucidate the effect of transient OGD on SUMO conjugation of target Proteins, we exposed neuroblastoma B35 cells expressing HA-SUMO3 to transient OGD and used stable isotope labeling with amino acids in cell culture (SILAC) to quantify OGD-induced changes in levels of specific SUMOylated Proteins. Lysates from control and OGD-treated cells were mixed equally, and HA-tagged Proteins were immunoprecipitated and analyzed by 1D-SDS-PAGE-LC-MS/MS. We identified 188 putative SUMO3-Conjugated Proteins, including numerous transcription factors and coregulators, and PIAS2 and PIAS4 SUMO ligases, of which 22 were increased or decreased >±2-fold. In addition to SUMO3, the levels of protein-Conjugated SUMO1 and SUMO2, as well as ubiquitin, were all increased. Importantly, protein ubiquitination induced by OGD was completely blocked by gene silencing of SUMO2/3. Collectively, these results suggest several mechanisms for OGD-modulated SUMOylation, point to a number of signaling pathways that may be targets of SUMO-based signaling and recovery from ischemic stress, as well as demonstrate a tightly controlled crosstalk between the SUMO and ubiquitin conjugation pathways.
Teresa G Hastings - One of the best experts on this subject based on the ideXlab platform.
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proteomic identification of dopamine Conjugated Proteins from isolated rat brain mitochondria and sh sy5y cells
Neurobiology of Disease, 2009Co-Authors: Victor S Van Laar, Amanda J. Mishizen, Michael Cascio, Teresa G HastingsAbstract:Abstract Dopamine oxidation has been previously demonstrated to cause dysfunction in mitochondrial respiration and membrane permeability, possibly related to covalent modification of critical Proteins by the reactive dopamine quinone. However, specific mitochondrial protein targets have not been identified. In this study, we utilized proteomic techniques to identify Proteins directly Conjugated with 14 C-dopamine from isolated rat brain mitochondria exposed to radiolabeled dopamine quinone (150 μM) and differentiated SH-SY5Y cells treated with 14 C-dopamine (150 μM). We observed a subset of rat brain mitochondrial Proteins that were covalently modified by 14 C-dopamine, including chaperonin, ubiquinol-cytochrome c reductase core protein 1, glucose regulated protein 75/mitochondrial HSP70/mortalin, mitofilin, and mitochondrial creatine kinase. We also found the Parkinson's disease associated Proteins ubiquitin carboxy-terminal hydrolase L1 and DJ-1 to be covalently modified by dopamine in both brain mitochondrial preparations and SH-SY5Y cells. The susceptibility of the identified Proteins to covalent modification by dopamine may carry implications for their role in the vulnerability of dopaminergic neurons in Parkinson's disease pathogenesis.
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Proteomic identification of dopamine-Conjugated Proteins from isolated rat brain mitochondria and SH-SY5Y cells
Neurobiology of Disease, 2009Co-Authors: Victor S. Van Laar, Amanda J. Mishizen, Michael Cascio, Teresa G HastingsAbstract:Dopamine oxidation has been previously demonstrated to cause dysfunction in mitochondrial respiration and membrane permeability, possibly related to covalent modification of critical Proteins by the reactive dopamine quinone. However, specific mitochondrial protein targets have not been identified. In this study, we utilized proteomic techniques to identify Proteins directly Conjugated with 14C-dopamine from isolated rat brain mitochondria exposed to radiolabeled dopamine quinone (150 μM) and differentiated SH-SY5Y cells treated with 14C-dopamine (150 μM). We observed a subset of rat brain mitochondrial Proteins that were covalently modified by 14C-dopamine, including chaperonin, ubiquinol-cytochrome c reductase core protein 1, glucose regulated protein 75/mitochondrial HSP70/mortalin, mitofilin, and mitochondrial creatine kinase. We also found the Parkinson's disease associated Proteins ubiquitin carboxy-terminal hydrolase L1 and DJ-1 to be covalently modified by dopamine in both brain mitochondrial preparations and SH-SY5Y cells. The susceptibility of the identified Proteins to covalent modification by dopamine may carry implications for their role in the vulnerability of dopaminergic neurons in Parkinson's disease pathogenesis. © 2009 Elsevier Inc. All rights reserved.