The Experts below are selected from a list of 80574 Experts worldwide ranked by ideXlab platform
Jicun Ren - One of the best experts on this subject based on the ideXlab platform.
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Studying Homo-Oligomerization and Hetero-Oligomerization of MDMX and MDM2 Proteins in Single Living Cells by Using In Situ Fluorescence Correlation Spectroscopy.
Biochemistry, 2021Co-Authors: Xiangyi Huang, Chaoqing Dong, Jicun RenAbstract:Protein Oligomerization plays a very important role in many physiological processes. p53 acts as a key tumor suppressor by regulating cell cycle arrest, DNA repair, and apoptosis, and its antitumor activity is regulated by the hetero- and homo-Oligomerization of MDMX and MDM2 proteins. So far, some traditional methods have been utilized to study the Oligomerization of MDMX and MDM2 in vitro, but they have not clarified some controversial issues or whether the extracellular results can represent the intracellular results. Here, we put forward an in situ method for studying protein homo- and hetero-Oligomerization in single living cells by using fluorescence correlation spectroscopy. In this study, MDMX and MDM2 were labeled with fluorescent proteins using lentiviral transfection. Autocorrelation spectroscopy and cross-correlation spectroscopy methods were used to study the Oligomerization of MDMX and MDM2 in situ and the effect of regulation of MDMX Oligomerization on p53-MDMX interactions in single living cells. We observed the homo- and hetero-Oligomerization of MDMX and MDM2 in living cells. Meanwhile, the levels of the homo-oligomers of MDMX and MDM2 were increased due to the lack of hetero-Oligomerization. Finally, the binding affinity of MDMX for p53 was improved with an increase in the level of MDMX hetero-Oligomerization.
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analyses of p73 protein Oligomerization and p73 mdm2 interaction in single living cells using in situ single molecule spectroscopy
Analytical Chemistry, 2021Co-Authors: Xiangyi Huang, Chaoqing Dong, Jicun RenAbstract:Protein Oligomerization and protein-protein interaction are crucial to regulate protein functions and biological processes. p73 protein is a very important transcriptional factor and can promote apoptosis and cell cycle arrest, and its transcriptional activity is regulated by p73 Oligomerization and p73-MDM2 interaction. Although extracellular studies on p73 Oligomerization and p73-MDM2 interaction have been carried out, it is unclear how p73 Oligomerization and p73-MDM2 interaction occur in living cells. In our study, we described an in situ method for studying p73 Oligomerization and p73-MDM2 interaction in living cells by combining fluorescence cross-correlation spectroscopy with a fluorescent protein labeling technique. Lentiviral transfection was used to transfect cells with a plasmid for either p73 or MDM2, each fused to a different fluorescent protein. p73 Oligomerization was evaluated using brightness per particle, and the p73-MDM2 interaction was quantified using the cross-correlation value. We constructed a series of p73 mutants in three domains (transactivation domain, DNA binding domain, and Oligomerization domain) and MDM2 mutants. We systematically studied p73 Oligomerization and the effects of p73 Oligomerization and the p73 and MDM2 structures on the p73-MDM2 interaction in single living cells. We have found that the p73 protein can form oligomers and that the p73 structure changes in the Oligomerization domain significantly influence its Oligomerization. p73 Oligomerization and the structure changes significantly affect the p73-MDM2 interaction. Furthermore, the effects of inhibitors on p73 Oligomerization and p73-MDM2 interaction were studied.
Chunbo Lou - One of the best experts on this subject based on the ideXlab platform.
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engineering the ultrasensitive transcription factors by fusing a modular Oligomerization domain
ACS Synthetic Biology, 2018Co-Authors: Junran Hou, Weiqian Zeng, Yeqing Zong, Zehua Chen, Chensi Miao, Baojun Wang, Chunbo LouAbstract:The dimerization and high-order Oligomerization of transcription factors has endowed them with cooperative regulatory capabilities that play important roles in many cellular functions. However, such advanced regulatory capabilities have not been fully exploited in synthetic biology and genetic engineering. Here, we engineered a C-terminally fused Oligomerization domain to improve the cooperativity of transcription factors. First, we found that two of three designed Oligomerization domains significantly increased the cooperativity and ultrasensitivity of a transcription factor for the regulated promoter. Then, seven additional transcription factors were used to assess the modularity of the Oligomerization domains, and their ultrasensitivity was generally improved, as assessed by their Hill coefficients. Moreover, we also demonstrated that the allosteric capability of the ligand-responsive domain remained intact when fusing with the designed Oligomerization domain. As an example application, we showed that ...
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Engineering the Ultrasensitive Transcription Factors by Fusing a Modular Oligomerization Domain
2018Co-Authors: Junran Hou, Weiqian Zeng, Yeqing Zong, Zehua Chen, Chensi Miao, Baojun Wang, Chunbo LouAbstract:The dimerization and high-order Oligomerization of transcription factors has endowed them with cooperative regulatory capabilities that play important roles in many cellular functions. However, such advanced regulatory capabilities have not been fully exploited in synthetic biology and genetic engineering. Here, we engineered a C-terminally fused Oligomerization domain to improve the cooperativity of transcription factors. First, we found that two of three designed Oligomerization domains significantly increased the cooperativity and ultrasensitivity of a transcription factor for the regulated promoter. Then, seven additional transcription factors were used to assess the modularity of the Oligomerization domains, and their ultrasensitivity was generally improved, as assessed by their Hill coefficients. Moreover, we also demonstrated that the allosteric capability of the ligand-responsive domain remained intact when fusing with the designed Oligomerization domain. As an example application, we showed that the engineered ultrasensitive transcription factor could be used to significantly improve the performance of a “stripe-forming” gene circuit. We envision that the Oligomerization modules engineered in this study could act as a powerful tool to rapidly tune the underlying response profiles of synthetic gene circuits and metabolic pathway controllers
Xiangyi Huang - One of the best experts on this subject based on the ideXlab platform.
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Studying Homo-Oligomerization and Hetero-Oligomerization of MDMX and MDM2 Proteins in Single Living Cells by Using In Situ Fluorescence Correlation Spectroscopy.
Biochemistry, 2021Co-Authors: Xiangyi Huang, Chaoqing Dong, Jicun RenAbstract:Protein Oligomerization plays a very important role in many physiological processes. p53 acts as a key tumor suppressor by regulating cell cycle arrest, DNA repair, and apoptosis, and its antitumor activity is regulated by the hetero- and homo-Oligomerization of MDMX and MDM2 proteins. So far, some traditional methods have been utilized to study the Oligomerization of MDMX and MDM2 in vitro, but they have not clarified some controversial issues or whether the extracellular results can represent the intracellular results. Here, we put forward an in situ method for studying protein homo- and hetero-Oligomerization in single living cells by using fluorescence correlation spectroscopy. In this study, MDMX and MDM2 were labeled with fluorescent proteins using lentiviral transfection. Autocorrelation spectroscopy and cross-correlation spectroscopy methods were used to study the Oligomerization of MDMX and MDM2 in situ and the effect of regulation of MDMX Oligomerization on p53-MDMX interactions in single living cells. We observed the homo- and hetero-Oligomerization of MDMX and MDM2 in living cells. Meanwhile, the levels of the homo-oligomers of MDMX and MDM2 were increased due to the lack of hetero-Oligomerization. Finally, the binding affinity of MDMX for p53 was improved with an increase in the level of MDMX hetero-Oligomerization.
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analyses of p73 protein Oligomerization and p73 mdm2 interaction in single living cells using in situ single molecule spectroscopy
Analytical Chemistry, 2021Co-Authors: Xiangyi Huang, Chaoqing Dong, Jicun RenAbstract:Protein Oligomerization and protein-protein interaction are crucial to regulate protein functions and biological processes. p73 protein is a very important transcriptional factor and can promote apoptosis and cell cycle arrest, and its transcriptional activity is regulated by p73 Oligomerization and p73-MDM2 interaction. Although extracellular studies on p73 Oligomerization and p73-MDM2 interaction have been carried out, it is unclear how p73 Oligomerization and p73-MDM2 interaction occur in living cells. In our study, we described an in situ method for studying p73 Oligomerization and p73-MDM2 interaction in living cells by combining fluorescence cross-correlation spectroscopy with a fluorescent protein labeling technique. Lentiviral transfection was used to transfect cells with a plasmid for either p73 or MDM2, each fused to a different fluorescent protein. p73 Oligomerization was evaluated using brightness per particle, and the p73-MDM2 interaction was quantified using the cross-correlation value. We constructed a series of p73 mutants in three domains (transactivation domain, DNA binding domain, and Oligomerization domain) and MDM2 mutants. We systematically studied p73 Oligomerization and the effects of p73 Oligomerization and the p73 and MDM2 structures on the p73-MDM2 interaction in single living cells. We have found that the p73 protein can form oligomers and that the p73 structure changes in the Oligomerization domain significantly influence its Oligomerization. p73 Oligomerization and the structure changes significantly affect the p73-MDM2 interaction. Furthermore, the effects of inhibitors on p73 Oligomerization and p73-MDM2 interaction were studied.
Shinichiroh Saitoh - One of the best experts on this subject based on the ideXlab platform.
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Ligand-dependent Toll-like receptor 4 (TLR4)-Oligomerization is directly linked with TLR4-signaling
Journal of Endotoxin Research, 2004Co-Authors: Shinichiroh Saitoh, Sachiko Akashi, Takenao Yamada, Natsuko Tanimura, Fumi Matsumoto, Koichi Fukase, Shoichi Kusumoto, Atsushi Kosugi, Kensuke MiyakeAbstract:Toll-like receptor 4 (TLR4) and MD-2 recognize lipid A, the active moiety of microbial lipopolysaccharide (LPS). Little is known about mechanisms for LPS recognition by TLR4/MD-2. We here showed, by using in vitro transfectants, ligand-induced TLR4-Oligomerization, which required both membrane CD14 and MD-2. We previously reported that lipid IVa, a lipid A precursor, is agonistic on mouse TLR4/MD-2 but antagonistic on human TLR4/MD-2 and chimeric mouse TLR4/human MD-2. Lipid IVa triggered Oligomerization of mouse TLR4/MD-2 but not human TLR4/MD-2 or chimeric mouse TLR4/human MD-2. Further, lipid IVa inhibited lipid A-dependent Oligomerization of chimeric mouse TLR4/human MD-2. These results demonstrate that ligand-induced TLR4-Oligomerization is directly linked with TLR4-signaling and suggest that MD-2 has an important role in regulating TLR4-Oligomerization.
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lipid a antagonist lipid iva is distinct from lipid a in interaction with toll like receptor 4 tlr4 md 2 and ligand induced tlr4 Oligomerization
International Immunology, 2004Co-Authors: Shinichiroh Saitoh, Sachiko Akashi, Takenao Yamada, Natsuko Tanimura, Makiko Kobayashi, Kazunori Konno, Fumi Matsumoto, Koichi Fukase, Shoichi Kusumoto, Yoshinori NagaiAbstract:: Toll-like receptor 4 (TLR4) and MD-2 recognizes lipid A, the active moiety of microbial lipopolysaccharide (LPS). Little is known about mechanisms for LPS recognition by TLR4-MD-2. Here we show ligand-induced TLR4 Oligomerization, homotypic interaction of TLR4, which directly leads to TLR4 signaling. Since TLR4 Oligomerization normally occurred in the absence of the cytoplasmic portion of TLR4, TLR4 Oligomerization works upstream of TLR4 signaling. Lipid IVa, a lipid A precursor, is agonistic on mouse TLR4-MD-2 but turns antagonistic on chimeric mouse TLR4-human MD-2, demonstrating that the antagonistic activity of lipid IVa is determined by human MD-2. Binding studies with radioactive lipid A and lipid IVa revealed that lipid IVa is similar to lipid A in dose-dependent and saturable binding to mouse TLR4-human MD-2. Lipid IVa, however, did not induce TLR4 Oligomerization, and inhibited lipid A-dependent Oligomerization of mouse TLR4-human MD-2. Thus, lipid IVa binds mouse TLR4-human MD-2 but does not trigger TLR4 Oligomerization. Binding study further revealed that the antagonistic activity of lipid IVa correlates with augmented maximal binding to mouse TLR4-human MD-2, which was approximately 2-fold higher than lipid A. Taken together, lipid A antagonist lipid IVa is distinct from lipid A in binding to TLR4-MD-2 and in subsequent triggering of TLR4 Oligomerization. Given that the antagonistic activity of lipid IVa is determined by MD-2, MD-2 has an important role in a link between ligand interaction and TLR4 Oligomerization.
Chaoqing Dong - One of the best experts on this subject based on the ideXlab platform.
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Studying Homo-Oligomerization and Hetero-Oligomerization of MDMX and MDM2 Proteins in Single Living Cells by Using In Situ Fluorescence Correlation Spectroscopy.
Biochemistry, 2021Co-Authors: Xiangyi Huang, Chaoqing Dong, Jicun RenAbstract:Protein Oligomerization plays a very important role in many physiological processes. p53 acts as a key tumor suppressor by regulating cell cycle arrest, DNA repair, and apoptosis, and its antitumor activity is regulated by the hetero- and homo-Oligomerization of MDMX and MDM2 proteins. So far, some traditional methods have been utilized to study the Oligomerization of MDMX and MDM2 in vitro, but they have not clarified some controversial issues or whether the extracellular results can represent the intracellular results. Here, we put forward an in situ method for studying protein homo- and hetero-Oligomerization in single living cells by using fluorescence correlation spectroscopy. In this study, MDMX and MDM2 were labeled with fluorescent proteins using lentiviral transfection. Autocorrelation spectroscopy and cross-correlation spectroscopy methods were used to study the Oligomerization of MDMX and MDM2 in situ and the effect of regulation of MDMX Oligomerization on p53-MDMX interactions in single living cells. We observed the homo- and hetero-Oligomerization of MDMX and MDM2 in living cells. Meanwhile, the levels of the homo-oligomers of MDMX and MDM2 were increased due to the lack of hetero-Oligomerization. Finally, the binding affinity of MDMX for p53 was improved with an increase in the level of MDMX hetero-Oligomerization.
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analyses of p73 protein Oligomerization and p73 mdm2 interaction in single living cells using in situ single molecule spectroscopy
Analytical Chemistry, 2021Co-Authors: Xiangyi Huang, Chaoqing Dong, Jicun RenAbstract:Protein Oligomerization and protein-protein interaction are crucial to regulate protein functions and biological processes. p73 protein is a very important transcriptional factor and can promote apoptosis and cell cycle arrest, and its transcriptional activity is regulated by p73 Oligomerization and p73-MDM2 interaction. Although extracellular studies on p73 Oligomerization and p73-MDM2 interaction have been carried out, it is unclear how p73 Oligomerization and p73-MDM2 interaction occur in living cells. In our study, we described an in situ method for studying p73 Oligomerization and p73-MDM2 interaction in living cells by combining fluorescence cross-correlation spectroscopy with a fluorescent protein labeling technique. Lentiviral transfection was used to transfect cells with a plasmid for either p73 or MDM2, each fused to a different fluorescent protein. p73 Oligomerization was evaluated using brightness per particle, and the p73-MDM2 interaction was quantified using the cross-correlation value. We constructed a series of p73 mutants in three domains (transactivation domain, DNA binding domain, and Oligomerization domain) and MDM2 mutants. We systematically studied p73 Oligomerization and the effects of p73 Oligomerization and the p73 and MDM2 structures on the p73-MDM2 interaction in single living cells. We have found that the p73 protein can form oligomers and that the p73 structure changes in the Oligomerization domain significantly influence its Oligomerization. p73 Oligomerization and the structure changes significantly affect the p73-MDM2 interaction. Furthermore, the effects of inhibitors on p73 Oligomerization and p73-MDM2 interaction were studied.
