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QUICK identification and SPR validation of signal transducers and activators of transcription 3 (Stat3) interacting proteins
Zheng, Peng2; Zhong, Qiu3; Xiong, Qian2; Yang, Mingkun2; Zhang, Jia1,2; Li, Chongyang2; Bi, Li-Jun1; Ge, Feng2; Bi, LJ (reprint author), Chinese Acad Sci, Natl Lab Biomacromol, Inst Biophys, Beijing 100101, Peoples R China
2012-01-04
Source PublicationJOURNAL OF PROTEOMICS
ISSN1874-3919
Volume75Issue:3Pages:1055-1066
AbstractSignal transducers and activators of transcription 3 (Stat3) has been reported to be involved in the pathogenesis of various human diseases and is constitutively active in human multiple myeloma (MM) U266 cells. The Stat3-regulated mechanisms involved in these processes, however, are not fully defined. To further understand the regulation of Stat3 activity, we performed a systematic proteomic analysis of Stat3 interacting proteins in U266 cells. This analysis, termed quantitative immunoprecipitation combined with knockdown (QUICK), combines RNAi, stable isotope labeling with amino acids in cell culture (SILAC), immunoprecipitation, and quantitative MS. As a result, quantitative mass spectrometry analysis allowed us to distinguish specific Stat3 interacting proteins from background proteins and led to the identification of a total of 38 proteins. Three Stat3 interacting proteins - 14-3-3 zeta, PRKCB and Hsp90 - were further confirmed by reciprocal co-immunoprecipitations and surface plasmon resonance (SPR) analysis. Our results therefore not only uncover a number of Stat3 interacting proteins that possess a variety of cellular functions, but also provide new insight into the mechanisms that regulate Stat3 activity and function in MM cells. (C) 2011 Elsevier B.V. All rights reserved.; Signal transducers and activators of transcription 3 (Stat3) has been reported to be involved in the pathogenesis of various human diseases and is constitutively active in human multiple myeloma (MM) U266 cells. The Stat3-regulated mechanisms involved in these processes, however, are not fully defined. To further understand the regulation of Stat3 activity, we performed a systematic proteomic analysis of Stat3 interacting proteins in U266 cells. This analysis, termed quantitative immunoprecipitation combined with knockdown (QUICK), combines RNAi, stable isotope labeling with amino acids in cell culture (SILAC), immunoprecipitation, and quantitative MS. As a result, quantitative mass spectrometry analysis allowed us to distinguish specific Stat3 interacting proteins from background proteins and led to the identification of a total of 38 proteins. Three Stat3 interacting proteins - 14-3-3 zeta, PRKCB and Hsp90 - were further confirmed by reciprocal co-immunoprecipitations and surface plasmon resonance (SPR) analysis. Our results therefore not only uncover a number of Stat3 interacting proteins that possess a variety of cellular functions, but also provide new insight into the mechanisms that regulate Stat3 activity and function in MM cells. (C) 2011 Elsevier B.V. All rights reserved.
SubtypeArticle
KeywordQuantitative Immunoprecipitation Combined With Knockdown (Quick) Stable Isotope Labeling With Amino Acids In Cell Culture (Silac) Stat3 14-3-3 Zeta Multiple Myeloma Surface Plasmon Resonance (Spr)
Department[Zhang, Jia; Bi, Li-Jun] Chinese Acad Sci, Natl Lab Biomacromol, Inst Biophys, Beijing 100101, Peoples R China; [Zheng, Peng; Xiong, Qian; Yang, Mingkun; Zhang, Jia; Li, Chongyang; Ge, Feng] Chinese Acad Sci, Inst Hydrobiol, Wuhan 430072, Peoples R China; [Zhong, Qiu] AntiTB Res Inst Guangdong Prov, IBP ARI Joint Ctr Res TB, Guangzhou 510630, Guangdong, Peoples R China
DOI10.1016/j.jprot.2011.10.020
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
Funding OrganizationChinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules ; Chinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules ; Chinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules ; Chinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules
Indexed BySCI
Language英语
WOS Research AreaBiochemistry & Molecular Biology
WOS SubjectBiochemical Research Methods
WOS IDWOS:000299719900027
WOS KeywordMULTIPLE-MYELOMA CELLS ; CONSTITUTIVE ACTIVATION ; PHOSPHATASE 2A ; KINASE-C ; IN-VIVO ; PROSTATE-CANCER ; KNOCKDOWN QUICK ; GENE ONTOLOGY ; TARGETING PKC ; APOPTOSIS
Funding OrganizationChinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules ; Chinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules ; Chinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules ; Chinese Academy of Sciences; National Basic Research Program of China (973 Program)[2012CB518700]; National Laboratory of Biomacromolecules
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Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/16763
Collection水生生物分子与细胞生物学研究中心_期刊论文
Corresponding AuthorBi, LJ (reprint author), Chinese Acad Sci, Natl Lab Biomacromol, Inst Biophys, Beijing 100101, Peoples R China
Affiliation1.Chinese Acad Sci, Natl Lab Biomacromol, Inst Biophys, Beijing 100101, Peoples R China
2.Chinese Acad Sci, Inst Hydrobiol, Wuhan 430072, Peoples R China
3.AntiTB Res Inst Guangdong Prov, IBP ARI Joint Ctr Res TB, Guangzhou 510630, Guangdong, Peoples R China
Recommended Citation
GB/T 7714
Zheng, Peng,Zhong, Qiu,Xiong, Qian,et al. QUICK identification and SPR validation of signal transducers and activators of transcription 3 (Stat3) interacting proteins[J]. JOURNAL OF PROTEOMICS,2012,75(3):1055-1066.
APA Zheng, Peng.,Zhong, Qiu.,Xiong, Qian.,Yang, Mingkun.,Zhang, Jia.,...&Bi, LJ .(2012).QUICK identification and SPR validation of signal transducers and activators of transcription 3 (Stat3) interacting proteins.JOURNAL OF PROTEOMICS,75(3),1055-1066.
MLA Zheng, Peng,et al."QUICK identification and SPR validation of signal transducers and activators of transcription 3 (Stat3) interacting proteins".JOURNAL OF PROTEOMICS 75.3(2012):1055-1066.
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