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Autophagosomal Proteome Analysis by Protein Correlation Profiling-SILAC

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Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC)

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1188))

Abstract

Autophagy is one of the two major degradation pathways within eukaryotic cells. Nevertheless, little is known about the protein composition of autophagosomes, the vesicles shuttling proteins to lysosomes for degradation. Protein correlation profiling in combination with stable isotope labeling by amino acids in cell culture is a stringent method to investigate the dynamics of the autophagosomal proteome. It enables the discrimination between autophagosomal and co-purifying proteins identifying organellar candidate proteins for further investigation.

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References

  1. Zimmermann AC, Zarei M, Eiselein S et al (2010) Quantitative proteomics for the analysis of spatio-temporal protein dynamics during autophagy. Autophagy 6:1009–1006

    Article  CAS  PubMed  Google Scholar 

  2. Engelke R, Becker AC, Dengjel J (2012) The degradative inventory of the cell: proteomic insights. Antioxid Redox Signal 17: 803–812

    Article  CAS  PubMed  Google Scholar 

  3. Dengjel J, Kratchmarova I, Blagoev B (2010) Mapping protein-protein interactions by quantitative proteomics. Methods Mol Biol 658: 267–278

    Article  CAS  PubMed  Google Scholar 

  4. Dengjel J, Kratchmarova I, Blagoev B (2009) Receptor tyrosine kinase signaling: a view from quantitative proteomics. Mol Biosyst 5: 1112–1121

    Article  CAS  PubMed  Google Scholar 

  5. Andersen JS, Wilkinson CJ, Mayor T et al (2003) Proteomic characterization of the human centrosome by protein correlation profiling. Nature 426:570–574

    Article  CAS  PubMed  Google Scholar 

  6. Dunkley TP, Hester S, Shadforth IP et al (2006) Mapping the Arabidopsis organelle proteome. Proc Natl Acad Sci U S A 103: 6518–6523

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Dunkley TP, Watson R, Griffin JL et al (2004) Localization of organelle proteins by isotope tagging (LOPIT). Mol Cell Proteomics 3: 1128–1134

    Article  CAS  PubMed  Google Scholar 

  8. Sadowski PG, Groen AJ, Dupree P et al (2008) Subcellular localization of membrane proteins. Proteomics 8:3991–4011

    Article  CAS  PubMed  Google Scholar 

  9. Dengjel J, Jakobsen L, Andersen JS (2010) Organelle proteomics by label-free and SILAC-based protein correlation profiling. Methods Mol Biol 658:255–265

    Article  CAS  PubMed  Google Scholar 

  10. Jakobsen L, Vanselow K, Skogs M et al (2011) Novel asymmetrically localizing components of human centrosomes identified by complementary proteomics methods. EMBO J 30: 1520–1535

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  11. Dengjel J, Hoyer-Hansen M, Nielsen MO et al (2012) Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens. Mol Cell Proteomics 11:M111.014035

    PubMed  Google Scholar 

  12. Shevchenko A, Tomas H, Havlis J et al (2006) In-gel digestion for mass spectrometric characterization of proteins and proteomes. Nat Protoc 1:2856–2860

    Article  CAS  PubMed  Google Scholar 

  13. Rappsilber J, Mann M, Ishihama Y (2007) Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips. Nat Protoc 2: 1896–1906

    Article  CAS  PubMed  Google Scholar 

  14. Cox J, Mann M (2008) MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification. Nat Biotechnol 26:1367–1372

    Article  CAS  PubMed  Google Scholar 

  15. Futschik ME, Carlisle B (2005) Noise-robust soft clustering of gene expression time-course data. J Bioinform Comput Biol 3:965–988

    Article  CAS  PubMed  Google Scholar 

  16. Rigbolt KT, Vanselow JT, Blagoev B (2011) GProX, a user-friendly platform for bioinformatics analysis and visualization of quantitative proteomics data. Mol Cell Proteomics 10: O110.007450

    PubMed  Google Scholar 

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Acknowledgments

The research leading to these results has received funding from the Excellence Initiative of the German Federal and State Governments through FRIAS and BIOSS, from the Deutsche Forschungsgemeinschaft (GZ DE1757/2-1), and from the Federal Ministry of Education and Research through GerontoSys II—NephAge (031 5896 A).

Author information

Authors and Affiliations

  1. Department of Dermatology, Medical Center - University of Freiburg, Hauptstr. 7, 79104, Freiburg, Germany

    Andrea C. Becker & Jörn Dengjel

  2. Freiburg Institute for Advanced Studies (FRIAS), University of Freiburg, Albertstr. 19, 79104, Freiburg, Germany

    Andrea C. Becker & Jörn Dengjel

  3. ZBSA Center for Biological Systems Analysis, University of Freiburg, Habsburgerstr. 49, 79104, Freiburg, Germany

    Andrea C. Becker & Jörn Dengjel

  4. BIOSS Centre for Biological Signalling Studies, University of Freiburg, Schänzlestr. 18, 79104, Freiburg, Germany

    Jörn Dengjel

Authors
  1. Andrea C. Becker
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  2. Jörn Dengjel
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Corresponding author

Correspondence to Jörn Dengjel .

Editor information

Editors and Affiliations

  1. Institute of Biology II, University of Freiburg, Freiburg, Germany

    Bettina Warscheid

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Becker, A.C., Dengjel, J. (2014). Autophagosomal Proteome Analysis by Protein Correlation Profiling-SILAC. In: Warscheid, B. (eds) Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC). Methods in Molecular Biology, vol 1188. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1142-4_19

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  • DOI: https://doi.org/10.1007/978-1-4939-1142-4_19

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1141-7

  • Online ISBN: 978-1-4939-1142-4

  • eBook Packages: Springer Protocols

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