Publications by authors named "Zhensheng Xie"

30 Publications

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Proteomics reveals the function reverse of MPSSS-treated prostate cancer-associated fibroblasts to suppress PC-3 cell viability via the FoxO pathway.

Cancer Med 2021 04 11;10(7):2509-2522. Epub 2021 Mar 11.

Key Laboratory of Protein and Peptide Pharmaceuticals & Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Prostate cancer-associated fibroblasts (prostate CAFs) are essential components of the tumor microenvironment and can promote tumor progression through their immunosuppressive functions. MPSSS, a novel polysaccharide purified from Lentinus edodes, has been reported to have anti-tumor activity. MPSSS could also inhibit the immunosuppressive function of prostate CAFs, which has been demonstrated through that the secretome of MPSSS-treated prostate CAFs could inhibit the proliferation of T cells. However, how the secretome of MPSSS-treated prostate CAFs influence prostate cancer progression is still unclear. Interestingly, we found that the low molecular weight (3-100kD) secretome of prostate CAFs (lmwCAFS) could promote the growth of PC-3 cells, while that of MPSSS-treated prostate CAFs (MT-lmwCAFS) could inhibit their growth. We carried out comparative secretomic analysis of lmwCAFS and MT-lmwCAFS to identify functional molecules that inhibit the growth of PC-3 cells, and proteomic analysis of lmwCAFS-treated PC-3 cells and MT-lmwCAFS-treated PC-3 cells to investigate the underlying molecular mechanism. These analyses suggest that TGF-β3 from MT-lmwCAFS may inhibit the growth of PC-3 cells. The validated experiments revealed that TGF-β3 from MT-lmwCAFS activated p21 expression in PC-3 cells by regulating the FoxO pathway thereby inducing G0/G1 cell cycle arrest of PC-3 cells. Overall, our data demonstrated that MPSSS reversed the ability of prostate CAFs to suppress the cell viability of PC-3 cells, which might provide a potential therapeutic strategy to prevent prostate cancer progression.
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http://dx.doi.org/10.1002/cam4.3825DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7982613PMC
April 2021

Oleic Acid and Eicosapentaenoic Acid Reverse Palmitic Acid-induced Insulin Resistance in Human HepG2 Cells via the Reactive Oxygen Species / JUN Pathway.

Genomics Proteomics Bioinformatics 2021 Feb 22. Epub 2021 Feb 22.

Key Laboratory of Protein and Peptide Pharmaceuticals & Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China. Electronic address:

The monounsaturated fatty acid (MUFA) oleic acid (OA) has previously been shown to reverse saturated fatty acid palmitic acid (PA)-induced hepatic insulin resistance (IR). However, its underlying molecular mechanism is unclear. In addition, previous studies have also shown that the ω-3 polyunsaturated fatty acid (PUFA) eicosapentaenoic acid (EPA) reverses PA-induced muscle IR, but whether EPA plays the same role in hepatic IR and its possible mechanism involved need to be further clarified. Here, we confirmed that EPA reversed PA-induced IR in HepG2 cells and compared the proteomic changes after treatment with different free fatty acids (FFAs). A total of 234 proteins were determined to be differentially expressed after MUFA treatment. Their functions were mainly related to responses to stress and endogenous stimuli, lipid metabolic process, and protein binding. For PUFA treatment, the PA-induced expression changes of 1326 proteins could be reversed by EPA treatment, 415 of which were mitochondrial proteins, with most of the functional proteins involved in oxidative phosphorylation (OXPHOS) and tricarboxylic acid (TCA) cycle. Mechanistic studies revealed that the protein encoded by JUN and reactive oxygen species (ROS) play a role in OA- and EPA-reversed PA-induced IR, respectively. EPA or OA alleviated PA-induced abnormal adenosine triphosphate (ATP) production, ROS generation, and calcium (Ca) content. Importantly, HO-activated production of ROS increased the protein expression of JUN, further resulting in IR in HepG2 cells. Taken together, we demonstrate that ROS/JUN is a common response pathway employed by HepG2 cells toward FFA-regulated IR.
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http://dx.doi.org/10.1016/j.gpb.2019.06.005DOI Listing
February 2021

Separation and characterization of extracellular vesicles from human plasma by asymmetrical flow field-flow fractionation.

Anal Chim Acta 2020 Aug 11;1127:234-245. Epub 2020 Jul 11.

Laboratory of Protein and Peptide Pharmaceuticals & Proteomics Laboratory, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China; University of Chinese Academy of Sciences, Beijing, 100049, China. Electronic address:

It is a big challenge to isolate extracellular vesicles (EVs) from human plasma because of the contamination from high abundant lipoproteins, such as high density lipoprotein (HDL) and low density lipoprotein particles (LDL). In this study, the parameters of asymmetrical flow field-flow fractionation (AF4) technology and sample preparation, including cross flow gradient, focusing time, ultrafiltration condition, sample amount and injection volume have been optimized and successfully utilized for the separation and characterization of EVs from human plasma. This study demonstrated that the great potential of AF4 in the separation of EVs from HDL and LDL in human plasma with high reproducibility and purity. This study indicated excessive focusing time in the AF4 separation and 100-300 kDa MWCO membrane based ultrafiltration in the pre-preparation will cause loss of EVs. A total of 1038 proteins have been identified in seven replicates of purified EVs from pooled human plasma sample. They are mainly enriched in extracellular exosomes, involved in extracellular matrix structural constituent, and associated with extracellular matrix-receptor interaction pathway. This study also indicated that human plasma contains more EVs than the paired serum at the same volume, and showed age- and gender-independent individual variability of the amount of EVs in human plasma. This study displayed that AF4 technique can serve as a powerful platform for the separation of EVs from human plasma, serum or human body fluids and this technology will promote the studies on EVs, such as proteomics, biomarker discovery and functions.
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http://dx.doi.org/10.1016/j.aca.2020.06.071DOI Listing
August 2020

Sequential Precipitation and Delipidation Enables Efficient Enrichment of Low-Molecular Weight Proteins and Peptides from Human Plasma.

J Proteome Res 2020 08 3;19(8):3340-3351. Epub 2020 Jul 3.

Key Laboratory of Protein and Peptide Pharmaceuticals & Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.

Low-molecular weight proteins and peptides (LMWPs, <30 kDa) in human plasma serve as potential biomarkers or drug targets and are endowed with desirable traits for biological and clinical studies. However, the identification of LMWPs from plasma is retarded by high-abundance proteins, high-molecular weight proteins, and lipids. Here, we present a sequential precipitation and delipidation (SPD) method for the efficient enrichment of LMWPs based on methyl--butyl ether/methanol/water systems. The enriched LMWP sample was analyzed by single-shot liquid chromatography-tandem mass spectrometry employing both HCD and EThcD without tryptic digestion, and 725 peptides were identified on average. The LMWP sample was also digested and analyzed using a bottom-up proteomics pipeline, and 289 proteins were identified, of which 129 (44.6%) proteins were less than 30 kDa and lipoprotein-associated proteins were significantly enriched. Additionally, 25 neuropeptides and 19 long noncoding RNA-encoded polypeptides were identified. Taken together, the SPD method shows good sensitivity and reproducibility when compared with other enrichment methods and has great potential for clinical biomarker discovery and application.
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http://dx.doi.org/10.1021/acs.jproteome.0c00232DOI Listing
August 2020

Proteomic Changes of Klebsiella pneumoniae in Response to Colistin Treatment and Mutation-Mediated Colistin Resistance.

Antimicrob Agents Chemother 2020 05 21;64(6). Epub 2020 May 21.

Key Laboratory of Protein and Peptide Pharmaceuticals and Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China

Polymyxins are increasingly used as the critical last-resort therapeutic options for multidrug-resistant Gram-negative bacteria. Unfortunately, polymyxin resistance has increased gradually over the past few years. Although studies on polymyxin mechanisms are expanding, systemwide analyses of the underlying mechanism for polymyxin resistance and stress response are still lacking. To understand how adapts to colistin (polymyxin E) pressure, we carried out proteomic analysis of a strain cultured with different concentrations of colistin. Our results showed that the proteomic responses to colistin treatment in involve several pathways, including (i) gluconeogenesis and the tricarboxylic acid (TCA) cycle, (ii) arginine biosynthesis, (iii) porphyrin and chlorophyll metabolism, and (iv) enterobactin biosynthesis. Interestingly, decreased abundances of class A β-lactamases, including TEM, SHV-11, and SHV-4, were observed in cells treated with colistin. Moreover, we present comprehensive proteome atlases of paired polymyxin-susceptible and -resistant strains. The polymyxin-resistant strain Ci, a mutant of ATCC BAA 2146, showed a missense mutation in This mutant, which displayed lipid A modification with 4-amino-4-deoxy-l-arabinose (l-Ara4N) and palmitoylation, showed striking increases in the expression of CrrAB, PmrAB, PhoPQ, ArnBCADT, and PagP. We hypothesize that mutations induce elevated expression of the operon and via PmrAB and PhoPQ. Moreover, the multidrug efflux pump KexD, which was induced by mutation, also contributed to colistin resistance. Overall, our results demonstrated proteomic responses to colistin treatment and the mechanism of CrrB-mediated colistin resistance, which may offer valuable information on the management of polymyxin resistance.
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http://dx.doi.org/10.1128/AAC.02200-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7269499PMC
May 2020

Comprehensive characterization and proteoform analysis of the hydrophobic surfactant proteins B and C in calf pulmonary surfactant.

J Pharm Biomed Anal 2019 Sep 21;174:625-632. Epub 2019 Jun 21.

Key Laboratory of Protein and Peptide Pharmaceuticals & Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China. Electronic address:

Calf pulmonary surfactant (CPS), which contains about 98% lipids and 2% hydrophobic surfactant proteins B (SP-B) and C (SP-C), has been used as a surfactant preparation for the clinical replacement therapy of respiratory distress syndrome (RDS). Characterization of SP-B and SP-C in CPS is informative for quality control and the evaluation of their biological activities. However, analysis of SP-B and SP-C is impeded by the high content of lipids in CPS. Here, we describe an integrated method by combining size exclusion chromatography (SEC)-based delipidation, SDS-PAGE separation, in-gel digestion and mass spectrometric analysis for comprehensive characterization and proteoform analysis of the extremely hydrophobic SP-B and SP-C in CPS. This study has shown that 30 proteoforms of SP-C with different truncations and modifications were identified and SP-B was found to be existed as a dimer form in the CPS.
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http://dx.doi.org/10.1016/j.jpba.2019.06.027DOI Listing
September 2019

Stepwise assembly of the earliest precursors of large ribosomal subunits in yeast.

Nucleic Acids Res 2017 Jun;45(11):6837-6847

Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.

Small ribosomal subunits are co-transcriptionally assembled on the nascent precursor rRNA in Saccharomyces cerevisiae. It is unknown how the highly intertwined structure of 60S large ribosomal subunits is initially formed. Here, we affinity purified and analyzed a series of pre-60S particles assembled in vivo on plasmid-encoded pre-rRNA fragments of increasing lengths, revealing a spatiotemporal assembly map for 34 trans-acting assembly factors (AFs), 30 ribosomal proteins and 5S rRNA. The gradual association of AFs and ribosomal proteins with the pre-rRNA fragments strongly supports that the pre-60S is co-transcriptionally, rather than post-transcriptionally, assembled. The internal and external transcribed spacers ITS1, ITS2 and 3΄ ETS in pre-rRNA must be processed in pre-60S. We show that the processing machineries for ITS1 and ITS2 are primarily recruited by the 5΄ and 3΄ halves of pre-27S RNA, respectively. Nevertheless, processing of both ITS1 and ITS2 requires a complete 25S region. The 3΄ ETS plays a minor role in ribosome assembly, but is important for efficient rRNA processing and ribosome maturation. We also identified a distinct pre-60S state occurring before ITS2 processing. Our data reveal the elusive co-transcriptional assembly pathway of large ribosomal subunit.
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http://dx.doi.org/10.1093/nar/gkx254DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5499802PMC
June 2017

Characterization and relative quantification of phospholipids based on methylation and stable isotopic labeling.

J Lipid Res 2016 Mar 5;57(3):388-97. Epub 2016 Jan 5.

Laboratory of Protein and Peptide Pharmaceuticals and Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China

Phospholipids (PLs), one of the lipid categories, are not only the primary building blocks of cellular membranes, but also can be split to produce products that function as second messengers in signal transduction and play a pivotal role in numerous cellular processes, including cell growth, survival, and motility. Here, we present an integrated novel method that combines a fast and robust TMS-diazomethane-based phosphate derivatization and isotopic labeling strategy, which enables simultaneous profiling and relative quantification of PLs from biological samples. Our results showed that phosphate methylation allows fast and sensitive identification of the six major PL classes, including their lysophospholipid counterparts, under positive ionization mode. The isotopic labeling of endogenous PLs was achieved by deuterated diazomethane, which was generated through acid-catalyzed hydrogen/deuterium (H/D) exchange and methanolysis of TMS-diazomethane during the process of phosphate derivatization. The measured H/D ratios of unlabeled and labeled PLs, which were mixed in known proportions, indicated that the isotopic labeling strategy is capable of providing relative quantitation with adequate accuracy, reproducibility, and a coefficient of variation of 9.1%, on average. This novel method offers unique advantages over existing approaches and presents a powerful tool for research of PL metabolism and signaling.
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http://dx.doi.org/10.1194/jlr.M063024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4766988PMC
March 2016

Proteomic Comparison and MRM-Based Comparative Analysis of Metabolites Reveal Metabolic Shift in Human Prostate Cancer Cell Lines.

J Proteome Res 2015 Aug 16;14(8):3390-402. Epub 2015 Jul 16.

§State Key Laboratory of Oncogenes and Related Genes, Renji-Med X Clinical Stem Cell Research Center, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127, China.

One of the major challenges in prostate cancer therapy remains the development of effective treatments for castration-resistant prostate cancer (CRPC), as the underlying mechanisms for its progression remain elusive. Previous studies showed that androgen receptor (AR) is crucially involved in regulation of metabolism in prostate cancer (PCa) cells throughout the transition from early stage, androgen-sensitive PCa to androgen-independent CRPC. AR achieves such metabolic rewiring directively either via its transcriptional activity or via interactions with AMP-activated protein kinase (AMPK). However, due to the heterogeneous expression and activity status of AR in PCa cells, it remains a challenge to investigate the links between AR status and metabolic alterations. To this end, we compared the proteomes of three pairs of androgen-sensitive (AS) and androgen-independent (AI) PCa cell lines, namely, PC3-AR(+)/PC3, 22Rv1/Du145, and LNCaP/C42B, using an iTRAQ labeling approach. Our results revealed that most of the differentially expressed proteins between each pair function in metabolism, indicating a metabolic shift between AS and AI cells, as further validated by multiple reaction monitoring (MRM)-based quantification of nucleotides and relative comparison of fatty acids between these cell lines. Furthermore, increased adenylate kinase isoenzyme 1 (AK1) in AS relative to AI cells may result in activation of AMPK, representing a major regulatory factor involved in the observed metabolic shift in PCa cells.
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http://dx.doi.org/10.1021/acs.jproteome.5b00464DOI Listing
August 2015

Dynamics of the lipid droplet proteome of the Oleaginous yeast rhodosporidium toruloides.

Eukaryot Cell 2015 Mar 9;14(3):252-64. Epub 2015 Jan 9.

Division of Biotechnology, Dalian Institute of Chemical Physics, CAS, Dalian, China Dalian National Laboratory for Clean Energy, Dalian Institute of Chemical Physics, CAS, Dalian, China

Lipid droplets (LDs) are ubiquitous organelles that serve as a neutral lipid reservoir and a hub for lipid metabolism. Manipulating LD formation, evolution, and mobilization in oleaginous species may lead to the production of fatty acid-derived biofuels and chemicals. However, key factors regulating LD dynamics remain poorly characterized. Here we purified the LDs and identified LD-associated proteins from cells of the lipid-producing yeast Rhodosporidium toruloides cultured under nutrient-rich, nitrogen-limited, and phosphorus-limited conditions. The LD proteome consisted of 226 proteins, many of which are involved in lipid metabolism and LD formation and evolution. Further analysis of our previous comparative transcriptome and proteome data sets indicated that the transcription level of 85 genes and protein abundance of 77 proteins changed under nutrient-limited conditions. Such changes were highly relevant to lipid accumulation and partially confirmed by reverse transcription-quantitative PCR. We demonstrated that the major LD structure protein Ldp1 is an LD marker protein being upregulated in lipid-rich cells. When overexpressed in Saccharomyces cerevisiae, Ldp1 localized on the LD surface and facilitated giant LD formation, suggesting that Ldp1 plays an important role in controlling LD dynamics. Our results significantly advance the understanding of the molecular basis of lipid overproduction and storage in oleaginous yeasts and will be valuable for the development of superior lipid producers.
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http://dx.doi.org/10.1128/EC.00141-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4346559PMC
March 2015

Screening and identification of potential biomarkers and establishment of the diagnostic serum proteomic model for the Traditional Chinese Medicine Syndromes of tuberculosis.

J Ethnopharmacol 2014 Sep 26;155(2):1322-31. Epub 2014 Jul 26.

Institute of Cell Biology, Zhejiang University, 388, Yuhangtang Road, Hangzhou 310058, PR China. Electronic address:

Ethnopharmacological Relevance: Chemotherapy is the mainstay of modern tuberculosis (TB) control. Traditional Chinese Medicine (TCM) can enhance the effect of anti-TB drug, promote the absorption of the foci in the lung and reduce drug toxicity. In TCM, the determination of treatment is based on ZHENG (also called TCM syndrome). To establish a diagnostic model by using proteomics technology in order to identify potential biomarkers for TCM syndromes of TB.

Materials And Methods: The surface-enhanced laser desorption ionization time of flight mass spectrometer (SELDI-TOF MS) combined with weak cation exchange (WCX) magnetic beads was used to screen serum samples from 71 cases of deficiency of lung yin syndrome (DLYS), 64 cases of fire (yang) excess yin deficiency syndrome (FEYDS) and 45 cases of deficiency of both qi and yin syndrome (DQYS). A classification model was established by Biomarker Pattern Software (BPS). Candidate protein biomarkers were purified by reverse phase-high performance liquid chromatograph (RP-HPLC), identified by MALDI-TOF MS, LC-MS/MS and validated by ProteinChip Immunoassays.

Results: A total of 74 discriminating m/z peaks (P<0.001) among three TCM syndromes of TB were detected. A diagnostic model for the TCM syndrome of TB based on the five biomarkers (3961.7, 4679.7, 5646.4, 8891.2 and 9416.7 m/z) was established which could discriminate DLYS, FEYDS and DQYS patients with an accuracy of 74.0%, 72.5%, and 96.7%, respectively. The candidate biomarker with m/z of 9416.7 was identified as a fragment of apolipoprotein C-III (apoC-III) by MALDI-TOF-MS and LC-MS/MS.

Conclusion: The TCM syndrome diagnostic model of TB could successfully distinguish the three TCM syndromes of TB patients. This provided a biological basis for the determination of treatment based on different TCM syndromes of TB. ApoC-III was identified as a potential biomarker for TCM syndromes of TB and revealed the biochemical basis and pathogenesis of TCM syndromes in TB.
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http://dx.doi.org/10.1016/j.jep.2014.07.025DOI Listing
September 2014

Integrated omics study delineates the dynamics of lipid droplets in Rhodococcus opacus PD630.

Nucleic Acids Res 2014 Jan 22;42(2):1052-64. Epub 2013 Oct 22.

National Laboratory of Macromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China, MOE Key Laboratory of Bioinformatics and Bioinformatics Division, Center for Synthetic and Systems Biology, TNLIST/Department of Automation, Tsinghua University, Beijing 100084, China, University of the Chinese Academy of Sciences, Beijing 100049, China, Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China, Department of Histology and Embryology, University of South China, Hengyang Hunan Province 421001, China, State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China, School of Applied Mathematics, Central University of Finance and Economics, Beijing 102206, China, Department of Molecular and Cell Biology, Center for Systems Biology, The University of Texas at Dallas, Dallas, TX 75083-0688, USA, Institut für Molekulare Mikrobiologie und Biotechnologie, Westfälische Wilhelms-Universität, Corrensstrasse 3, D-48149 Münster, Germany, Environmental Sciences Department, King Abdulaziz University, Jeddah 21589, Saudi Arabia, Department of Anatomy and Molecular Cell Biology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa, Nagoya 466-8550, Japan and School of Life Sciences, La Trobe University, Melbourne, Victoria, 3086, Australia.

Rhodococcus opacus strain PD630 (R. opacus PD630), is an oleaginous bacterium, and also is one of few prokaryotic organisms that contain lipid droplets (LDs). LD is an important organelle for lipid storage but also intercellular communication regarding energy metabolism, and yet is a poorly understood cellular organelle. To understand the dynamics of LD using a simple model organism, we conducted a series of comprehensive omics studies of R. opacus PD630 including complete genome, transcriptome and proteome analysis. The genome of R. opacus PD630 encodes 8947 genes that are significantly enriched in the lipid transport, synthesis and metabolic, indicating a super ability of carbon source biosynthesis and catabolism. The comparative transcriptome analysis from three culture conditions revealed the landscape of gene-altered expressions responsible for lipid accumulation. The LD proteomes further identified the proteins that mediate lipid synthesis, storage and other biological functions. Integrating these three omics uncovered 177 proteins that may be involved in lipid metabolism and LD dynamics. A LD structure-like protein LPD06283 was further verified to affect the LD morphology. Our omics studies provide not only a first integrated omics study of prokaryotic LD organelle, but also a systematic platform for facilitating further prokaryotic LD research and biofuel development.
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http://dx.doi.org/10.1093/nar/gkt932DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3902926PMC
January 2014

Proteomic studies of isolated lipid droplets from bacteria, C. elegans, and mammals.

Methods Cell Biol 2013 ;116:1-14

National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China; University of Chinese Academy of Sciences, Beijing, China.

Lipid droplets (LDs) are an intracellular organelle, consisting of a neutral lipid core covered by a monolayer of phospholipids and proteins. It primarily mediates lipid storage, metabolism, and transportation. Recently, research of LDs has emerged as a rapidly developing field due to the strong linkage between ectopic lipid accumulation and metabolic syndromes. Recently, more than 30 proteomic studies of isolated LDs have identified many important LD proteins that have highlighted and have also predicted the potential biological roles of the organelle, motivating the field to develop quite rapidly. This chapter summarizes methods used in proteomic studies for three representative species reported and discusses their advantages and disadvantages. We believe that this chapter provides useful information and methods for future LD proteomic studies especially for LDs in other species.
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http://dx.doi.org/10.1016/B978-0-12-408051-5.00001-2DOI Listing
May 2014

Chronic high glucose induced INS-1β cell mitochondrial dysfunction: a comparative mitochondrial proteome with SILAC.

Proteomics 2013 Oct;13(20):3030-9

Laboratory of Protein and Peptide Pharmaceuticals, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China; Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

As glucose-stimulated insulin secretion of pancreatic β cell is triggered and promoted by the metabolic messengers derived from mitochondria, mitochondria take a central stage in the normal function of β cells. β cells in diabetics were chronically exposed to hyperglycemia stimulation, which have been reported to exert deleterious effects on β-cell mitochondria. However, the mechanism of the toxic effects of hyperglycemia on β-cell mitochondria was not clear. In this study, we characterized the biological functional changes of rat INS-1β cells and their mitochondria with chronic exposure to hyperglycemia and created a research model of chronic hyperglycemia-induced dysfunctional β cells with damaged mitochondria. Then, SILAC-based quantitative proteomic approach was used to compare the mitochondrial protein expression from high glucose treated INS-1β cells and control cells. The expression of some mitochondrial proteins was found with significant changes. Functional classification revealed most of these proteins were related with oxidative phosphorylation, mitochondrial protein biosynthesis, substances metabolism, transport, and cell death. These results presented some useful information about the effect of glucotoxicity on the β-cell mitochondria.
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http://dx.doi.org/10.1002/pmic.201200448DOI Listing
October 2013

Proteomic study and marker protein identification of Caenorhabditis elegans lipid droplets.

Mol Cell Proteomics 2012 Aug 9;11(8):317-28. Epub 2012 Apr 9.

Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.

Lipid droplets (LDs) are a neutral lipid storage organelle that is conserved across almost all species. Many metabolic syndromes are directly linked to the over-storage of neutral lipids in LDs. The study of LDs in Caenorhabditis elegans (C. elegans) has been difficult because of the lack of specific LD marker proteins. Here we report the purification and proteomic analysis of C. elegans lipid droplets for the first time. We identified 306 proteins, 63% of these proteins were previously known to be LD-proteins, suggesting a similarity between mammalian and C. elegans LDs. Using morphological and biochemical analyses, we show that short-chain dehydrogenase, DHS-3 is almost exclusively localized on C. elegans LDs, indicating that it can be used as a LD marker protein in C. elegans. These results will facilitate further mechanistic studies of LDs in this powerful genetic system, C. elegans.
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http://dx.doi.org/10.1074/mcp.M111.016345DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3412964PMC
August 2012

Purification, identification and profiling of serum amyloid A proteins from sera of advanced-stage cancer patients.

J Chromatogr B Analyt Technol Biomed Life Sci 2012 Mar 16;889-890:3-9. Epub 2012 Jan 16.

Laboratory of Protein and Peptide Pharmaceuticals & Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Surface-enhanced laser desorption/ionization time of flight mass spectrometry (SELDI-TOF-MS) is a powerful tool for screening potential biomarkers of various pathological conditions. However, low resolution and mass accuracy of SELDI-TOF-MS remain a major obstacle for determination of biological identities of potential protein biomarkers. We report here a refined workflow that combines ZipTip desalting, acetonitrile precipitation, high-performance liquid chromatography (HPLC) separation and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) analysis for the profiling, purification and identification of the targeted serum proteins found by SELDI-TOF-MS. By using this workflow, we purified ten targeted proteins from the sera of patients with various types of advanced stage (stage III-IV) cancers. These proteins were identified as isoforms of the human serum amyloid protein A (SAA) family with or without truncations at their N-terminals. This was confirmed by Western blot analysis. Different SAA expression patterns were observed by MALDI-TOF-MS profiling. SAA has long been reported as a biomarker for various cancer types such as lung cancer, ovarian cancer, and breast cancer. However, in this study we found increased SAA expression in the sera of advanced-stage cancer patients with different cancer types. Our results suggest that maybe SAA should not be used alone as a biomarker for any specific cancer type.
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http://dx.doi.org/10.1016/j.jchromb.2012.01.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7105184PMC
March 2012

Diagnostic serum proteomic analysis in patients with active tuberculosis.

Clin Chim Acta 2012 May 4;413(9-10):883-7. Epub 2012 Feb 4.

Army Tuberculosis Key Laboratory, Institute of Tuberculosis Research, the 309th Hospital of PLA, Beijing 100091, China.

Background: The diagnosis for smear-negative pulmonary tuberculosis (TB) is very difficult. Proteomic fingerprinting of sera is a potentially useful tool.

Methods: This study analyzed the results of the proteomic fingerprinting in sera obtained from active TB patients and controls using the surface-enhanced laser desorption ionization time of flight mass spectrometry (SELDI-TOF-MS) and protein-chip technology. The peaks were detected and analyzed, and a diagnostic system was developed. The protein peak was identified using high performance liquid chromatography (HPLC)-tandem matrix-assisted laser desorption/ionization-TOF-MS (MALDI-TOF-MS).

Results: Around 50 protein peaks were found significantly different between the TB patients and the controls (P<0.01). Three protein peaks at m/z 5643, 4486 and 4360 were selected for system classification and the development of a decision model. The model distinguished the TB patients from the controls with a sensitivity of 96.9% and a specificity of 97.8%, respectively. The diagnostic accuracy was up to 97.3%. The one most discrepant protein peak at m/z 5643 seen in sera of active TB patients was identified as orosomucoid.

Conclusions: A diagnostic system for active TB was developed using mass spectrometry and protein chip technology and required only small-volume serum samples. One potential protein biomarker at m/z 5643 was identified as orosomucoid.
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http://dx.doi.org/10.1016/j.cca.2012.01.036DOI Listing
May 2012

Bioinformatics analysis of proteomic profiles during the process of anti-Thy1 nephritis.

Mol Cell Proteomics 2012 Apr 7;11(4):M111.008755. Epub 2011 Dec 7.

Department of Nephrology, State Key Lab of Kidney Diseases, General Hospital of PLA, Beijing, China.

Anti-Thy1 nephritis is a well-established experimental mesangial proliferative nephritis model. Exploring the molecular mechanisms of pathophysiology in anti-Thy1 nephritis may elucidate the pathogeneses of mesangial proliferation. We examined the roles and acting mechanisms of differentially expressed proteins (DEPs) by bioinformatics analysis of glomeruli proteomic profiles during the course of anti-Thy1 nephritis. In total, 108 DEPs were found by two-dimensional fluorescence difference gel electrophoresis (2D-DIGE), and 40 DEPs were identified by matrix-assisted laser desorption ionization/time of flight and liquid chromatography-MS. DEPs were classified into five clusters (Clusters 1-5), according to their expression trends using Cluster 3.0 software, involved in regulating biological processes such as the stress response, cell proliferation, apoptosis, energy metabolism, transport, and the actin cytoskeleton. The expression patterns of ten DEPs, distributed across five clusters, including AKR1A1, AGAT, ATP6V1B2, HIBADH, MDH1, MPST, NIT2, PRDX6, PSMB7, and TPI1, were validated by Western blotting. Based on Western blotting and immunohistochemistry, we also found that the DEP FHL2, which was primarily expressed in the mesangial region, was down-regulated on days 3 and 5, and up-regulated on day 10. In vitro, we found that FHL2 overexpression induced mesangial cell proliferation by increasing the number of S-phase cells and decreasing G2/M-phase cells, whereas inhibiting FHL2 had the opposite effect. This study explored novel DEPs and their expression patterns during anti-Thy1 nephritis, and elucidated FHL2's effect on mesangial cell proliferation. These results will contribute to our understanding of the pathogenesis of mesangial proliferation.
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http://dx.doi.org/10.1074/mcp.M111.008755DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3322559PMC
April 2012

Quantitative proteomics analysis of parthenogenetically induced pluripotent stem cells.

Protein Cell 2011 Aug 9;2(8):631-46. Epub 2011 Sep 9.

Department of Cell Biology and Genetics, Key Laboratory of Bioactive Materials of Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China.

Parthenogenetic embryonic stem (pES) cells isolated from parthenogenetic activation of oocytes and embryos, also called parthenogenetically induced pluripotent stem cells, exhibit pluripotency evidenced by both in vitro and in vivo differentiation potential. Differential proteomic analysis was performed using differential in-gel electrophoresis and isotope-coded affinity tag-based quantitative proteomics to investigate the molecular mechanisms underlying the developmental pluripotency of pES cells and to compare the protein expression of pES cells generated from either the in vivo-matured ovulated (IVO) oocytes or from the in vitro-matured (IVM) oocytes with that of fertilized embryonic stem (fES) cells derived from fertilized embryos. A total of 76 proteins were upregulated and 16 proteins were downregulated in the IVM pES cells, whereas 91 proteins were upregulated and 9 were downregulated in the IVO pES cells based on a minimal 1.5-fold change as the cutoff value. No distinct pathways were found in the differentially expressed proteins except for those involved in metabolism and physiological processes. Notably, no differences were found in the protein expression of imprinted genes between the pES and fES cells, suggesting that genomic imprinting can be corrected in the pES cells at least at the early passages. The germline competent IVM pES cells may be applicable for germ cell renewal in aging ovaries if oocytes are retrieved at a younger age.
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http://dx.doi.org/10.1007/s13238-011-1081-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4875327PMC
August 2011

Preparative isolation of alkaloids from Corydalis bungeana Turcz. by high-speed counter-current chromatography using stepwise elution.

J Sep Sci 2011 May 8;34(9):987-94. Epub 2011 Mar 8.

Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, P R China.

High-speed counter-current chromatography (HSCCC) was successfully applied for the preparative separation and purification of alkaloids from Corydalis bungeana Turcz. (Kudiding in Chinese) for the first time. After the measurement of partition coefficient of seven target alkaloids in the nine two-phase solvent systems composed of CHCl(3)-MeOH-(0.1 M; 0.2 M; 0.3 M) HCl (4:1.5:2; 4:2:2; 4:3:2, v/v), CHCl(3)-MeOH-0.2 M HCl (4:2:2, v/v) and CHCl(3)-MeOH-0.3 M HCl (4:3:2, v/v) were finally selected for the HSCCC separation using the first upper phase as the stationary phase and the stepwise elution of the two lower mobile phases. Consequently, sanguinarine (10 mg), corynoline (25 mg), protopine (20 mg), corynoloxine (18 mg), and 12-hydroxycorynoline (8 mg) were obtained from 200 mg of crude alkaloid extracts with purities of 94-99% as determined by HPLC. Their chemical structures were characterized on the basis of (1)H-NMR, (13)C-NMR, and LC-ESI-Q-TOF-MS/MS analyses.
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http://dx.doi.org/10.1002/jssc.201000785DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3086934PMC
May 2011

Acyl-biotinyl exchange chemistry and mass spectrometry-based analysis of palmitoylation sites of in vitro palmitoylated rat brain tubulin.

Protein J 2010 Nov;29(8):531-7

Department of Basic Medical Sciences, Medical College of Taizhou University, Taizhou, Zhejiang 318000, China.

Research has shown that the palmitoyl group of α-tubulin mediates the hydrophobic interaction between microtubules and intracellular membranes and that palmitoylated tubulin plays a role in signal transduction. There are 20 cysteine residues per α/β tubulin heterodimer. C376 of α-tubulin was reported to be predominantly palmitoylated and C20, C213 and C305 of α-tubulin were palmitoylated at lower levels. The previous method used for the analysis of the palmitoylation sites on α-tubulin was based on ³H-labeling, enzymolysis, purification and sequencing. This approach, although efficient, is laborious. Mass spectrometry (MS), especially tandem MS, has been shown to be a successful method for identification of various post-translational modifications of proteins. We report here a convenient MS-based method to comprehensively analyze the palmitoylation sites of the α/β tubulin heterodimer. Acyl-biotinyl exchange chemistry and streptavidin agarose affinity purification were applied to enrich palmitoylated peptides from tubulin. After nano-LC-MS/MS analysis, database searching and manual analysis of the spectra revealed that 11 cysteine residues of the α/β tubulin heterodimer were palmitoylated.
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http://dx.doi.org/10.1007/s10930-010-9285-xDOI Listing
November 2010

Mammalian mitochondrial proteomics: insights into mitochondrial functions and mitochondria-related diseases.

Expert Rev Proteomics 2010 Jun;7(3):333-45

Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Mitochondria are organelles that are essential for cell life and death. A huge range of pathologies, including neurodegenerative diseases, cancer, diabetes and aging, have been reported to be associated with mitochondrial dysfunction. Therefore, identification of mitochondrial proteins that are differentially expressed in these pathologies will help to further our understanding of these diseases. In recent years, great achievements have been made in mammalian mitochondrial proteomics. Here we provide an overview of the current state of knowledge with respect to the whole mitochondrial proteome, the mitochondrial subproteome, mitochondrial complexes and mitochondrial post-translational modifications. Applications of comparative mitochondrial proteomics to various pathologies that have provided clues for understanding the relationship between mitochondrial dysfunction and pathogenesis are described. We conclude that mitochondrial proteomics can be used not only to map all the components of mitochondria, but can also provide information for discovering therapeutic targets for mitochondria-related diseases.
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http://dx.doi.org/10.1586/epr.10.22DOI Listing
June 2010

The profile of mitochondrial proteins and their phosphorylation signaling network in INS-1 beta cells.

J Proteome Res 2010 Jun;9(6):2898-908

Laborotary of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Mitochondria have important roles in cellular physiological functions and various diseases. In pancreatic beta cells, mitochondria play a central role in glucose-stimulated insulin secretion (GSIS). To reveal the potential functions of mitochondria in the GSIS process in beta cells, shotgun proteomics was applied to profiling mitochondrial proteins and their potential phosphorylation sites in rat INS-1 cells. More than 800 proteins were assigned to mitochondria. In addition, 84 different mitochondrial phosphoproteins were identified, and 52 upstream kinases of mitochondrial phosphoproteins were predicted using bioinformatics tools. Regulation networks of mitochondrial phosphoproteins were constructed by integrating mitochondrial protein interaction networks and mitochondrial phosphorylation signaling, providing a preliminary survey of how phosphorylation signaling regulates mitochondrial function in beta cells. We present integrated resources including the protein composition and signaling pathways of mitochondria which can be used to understand the role of mitochondria in GSIS.
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http://dx.doi.org/10.1021/pr100139zDOI Listing
June 2010

Enhanced MALDI-TOF MS analysis of phosphopeptides using an optimized DHAP/DAHC matrix.

J Biomed Biotechnol 2010 21;2010:759690. Epub 2010 Mar 21.

Proteomic Platform and National Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Selecting an appropriate matrix solution is one of the most effective means of increasing the ionization efficiency of phosphopeptides in matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). In this study, we systematically assessed matrix combinations of 2, 6-dihydroxyacetophenone (DHAP) and diammonium hydrogen citrate (DAHC), and demonstrated that the low ratio DHAP/DAHC matrix was more effective in enhancing the ionization of phosphopeptides. Low femtomole level of phosphopeptides from the tryptic digests of alpha-casein and beta-casein was readily detected by MALDI-TOF-MS in both positive and negative ion mode without desalination or phosphopeptide enrichment. Compared with the DHB/PA matrix, the optimized DHAP/DAHC matrix yielded superior sample homogeneity and higher phosphopeptide measurement sensitivity, particularly when multiple phosphorylated peptides were assessed. Finally, the DHAP/DAHC matrix was applied to identify phosphorylation sites from alpha-casein and beta-casein and to characterize two phosphorylation sites from the human histone H1 treated with Cyclin-Dependent Kinase-1 (CDK1) by MALDI-TOF/TOF MS.
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http://dx.doi.org/10.1155/2010/759690DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2842900PMC
June 2010

Phosphoproteome analysis of rat L6 myotubes using reversed-phase C18 prefractionation and titanium dioxide enrichment.

J Proteome Res 2010 Feb;9(2):777-88

Proteomic Platform, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

The rat L6 myotubes is an important in vitro model system for studying signaling pathways in skeletal muscle. Exploring phosphorylation events involved in the skeletal muscle is very significant for elucidating the kinase-substrate relationship, understanding regulatory mechanisms involved in signaling pathways and providing insights into numerous cell processes. Here, we used mass spectrometry-based proteomics to conduct global phosphoproteome profiling of rat L6 myotubes. Using an efficient phosphoproteomic strategy including prefractionation of tryptic peptide mixtures with self-packed RP C18 columns, phosphopeptide enrichment with TiO(2), and 2D-LC (SCX/RP)-MS/MS analysis, a total of 2230 unique phosphopeptides from 1195 proteins were identified with a false-discovery rate of less than 1.0% using a target/decoy database searching strategy. After determining the degree of certainty of the phosphorylation site location (Ascore value >or=19), 11 Ser motifs and one Thr motif were derived from our data set using the Motif-X algorithm. Several potential signaling pathways were found in our myotubes phosphoproteome, such as the MAPK signaling pathway and the IGF-1/Insulin signaling pathway.
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http://dx.doi.org/10.1021/pr900646kDOI Listing
February 2010

Proteomic analysis of mitochondria from Caenorhabditis elegans.

Proteomics 2009 Oct;9(19):4539-53

Proteomic Platform, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, PR China.

Mitochondria play essential roles in cell physiological processes including energy production, metabolism, ion homeostasis, cell growth, aging and apoptosis. Proteomic strategies have been applied to the study of mitochondria since 1998; these studies have yielded decisive information about the diverse physiological functions of the organelle. As an ideal model biological system, the nematode Caenorhabditis elegans has been widely used in the study of several diseases, such as metabolic diseases and cancer. However, the mitochondrial proteome of C. elegans remains elusive. In this study, we purified mitochondria from C. elegans and performed a comprehensive proteomic analysis using the shotgun proteomic approach. A total of 1117 proteins have been identified with at least two unique peptides. Their physicochemical and functional characteristics, subcellular locations, related biological processes, and associations with human diseases, especially Parkinson's disease, are discussed. An orthology comparison was also performed between C. elegans and four other model organisms for a general depiction of the conservation of mitochondrial proteins during evolution. This study will provide new clues for understanding the role of mitochondria in the physiological and pathological processes of C. elegans.
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http://dx.doi.org/10.1002/pmic.200900101DOI Listing
October 2009

Discovery and identification of potential biomarkers of pediatric acute lymphoblastic leukemia.

Proteome Sci 2009 Mar 16;7. Epub 2009 Mar 16.

Proteomic Platform, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, PR China.

Background: Acute lymphoblastic leukemia (ALL) is a common form of cancer in children. Currently, bone marrow biopsy is used for diagnosis. Noninvasive biomarkers for the early diagnosis of pediatric ALL are urgently needed. The aim of this study was to discover potential protein biomarkers for pediatric ALL.

Methods: Ninety-four pediatric ALL patients and 84 controls were randomly divided into a "training" set (45 ALL patients, 34 healthy controls) and a test set (49 ALL patients, 30 healthy controls and 30 pediatric acute myeloid leukemia (AML) patients). Serum proteomic profiles were measured using surface-enhanced laser desorption/ionization-time-of-flight mass spectroscopy (SELDI-TOF-MS). A classification model was established by Biomarker Pattern Software (BPS). Candidate protein biomarkers were purified by HPLC, identified by LC-MS/MS and validated using ProteinChip immunoassays.

Results: A total of 7 protein peaks (9290 m/z, 7769 m/z, 15110 m/z, 7564 m/z, 4469 m/z, 8937 m/z, 8137 m/z) were found with differential expression levels in the sera of pediatric ALL patients and controls using SELDI-TOF-MS and then analyzed by BPS to construct a classification model in the "training" set. The sensitivity and specificity of the model were found to be 91.8%, and 90.0%, respectively, in the test set. Two candidate protein peaks (7769 and 9290 m/z) were found to be down-regulated in ALL patients, where these were identified as platelet factor 4 (PF4) and pro-platelet basic protein precursor (PBP). Two other candidate protein peaks (8137 and 8937 m/z) were found up-regulated in the sera of ALL patients, and these were identified as fragments of the complement component 3a (C3a).

Conclusion: Platelet factor (PF4), connective tissue activating peptide III (CTAP-III) and two fragments of C3a may be potential protein biomarkers of pediatric ALL and used to distinguish pediatric ALL patients from healthy controls and pediatric AML patients. Further studies with additional populations or using pre-diagnostic sera are needed to confirm the importance of these findings as diagnostic markers of pediatric ALL.
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http://dx.doi.org/10.1186/1477-5956-7-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2662805PMC
March 2009

Preliminary quantitative profile of differential protein expression between rat L6 myoblasts and myotubes by stable isotope labeling with amino acids in cell culture.

Proteomics 2009 Mar;9(5):1274-92

Proteomics Platform, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Defining the mechanisms governing myogenesis has advanced in recent years. Skeletal-muscle differentiation is a multi-step process controlled spatially and temporally by various factors at the transcription level. To explore those factors involved in myogenesis, stable isotope labeling with amino acids in cell culture (SILAC), coupled with high-accuracy mass spectrometry (LTQ-Orbitrap), was applied successfully. Rat L6 cell line is an excellent model system for studying muscle myogenesis in vitro. When mononucleate L6 myoblast cells reach confluence in culture plate, they could transform into multinucleate myotubes by serum starvation. By comparing protein expression of L6 myoblasts and terminally differentiated multinucleated myotubes, 1170 proteins were quantified and 379 proteins changed significantly in fully differentiated myotubes in contrast to myoblasts. These differentially expressed proteins are mainly involved in inter-or intracellular signaling, protein synthesis and degradation, protein folding, cell adhesion and extracellular matrix, cell structure and motility, metabolism, substance transportation, etc. These findings were supported by many previous studies on myogenic differentiation, of which many up-regulated proteins were found to be involved in promoting skeletal muscle differentiation for the first time in our study. In summary, our results provide new clues for understanding the mechanism of myogenesis.
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http://dx.doi.org/10.1002/pmic.200800354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2946197PMC
March 2009

Rad9 plays an important role in DNA mismatch repair through physical interaction with MLH1.

Nucleic Acids Res 2008 Nov 8;36(20):6406-17. Epub 2008 Oct 8.

National Laboratory of Biomacromolecules, Center for Infection and Immunity, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

Rad9 is conserved from yeast to humans and plays roles in DNA repair (homologous recombination repair, and base-pair excision repair) and cell cycle checkpoint controls. It has not previously been reported whether Rad9 is involved in DNA mismatch repair (MMR). In this study, we have demonstrated that both human and mouse Rad9 interacts physically with the MMR protein MLH1. Disruption of the interaction by a single-point mutation in Rad9 leads to significantly reduced MMR activity. This disruption does not affect S/M checkpoint control and the first round of G(2)/M checkpoint control, nor does it alter cell sensitivity to UV light, gamma rays or hydroxyurea. Our data indicate that Rad9 is an important factor in MMR and carries out its MMR function specifically through interaction with MLH1.
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http://dx.doi.org/10.1093/nar/gkn686DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2582629PMC
November 2008

Pedigree analysis of an elite rice hybrid using proteomic approach.

Proteomics 2006 Jan;6(2):474-86

Beijing Genomics Institute, Chinese Academy of Sciences, Beijing, PR China.

The definition of dominance or epistasis is generally on the basis of a descriptive characterization for these crops in the field, such as yield per hectare and the weight of grain. Since these trait examinations lack molecular information, how to precisely predict the phenotypic changes in filial generation is still a problem in heterosis studies. For rice, the genetic information caused by hybridization can be archived through analyzing of proteomes of rice seeds. Differential analysis of proteomes was introduced for the rice seeds of three cultivars, 9311, PA64S and LYP9, an elite rice hybrid from cross between 9311 and PA64S. In the three rice endosperms, the expression profiles of proteins were similar with the stained spots of 47 +/- 1, 46 +/- 0.6 and 44 +/- 0.6, for 9311, PA64S and LYP9, respectively; however, the number of proteins expressed in the rice embryos was significantly increased with the stained spots of 395.3 +/- 12.9, 350 +/- 9.2, and 389.3 +/- 16.4, for 9311, PA64S and LYP9, respectively. Importantly, the image comparisons and protein identifications have revealed in significantly different embryo protein spots among the three rice cultivars. By carefully analyzing these different 2-DE spots, many of them from the three embryos were shown to display a mirrored relationships between parents and the first filial generation. Furthermore, all of stained spots in LYP9 embryo were found on the 2-DEs from its parents, indicating that there was a genetic linkage. These results suggest that proteomic approach is able to serve pedigree analysis and functional prediction for new rice breeds.
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http://dx.doi.org/10.1002/pmic.200500227DOI Listing
January 2006