Brief Communication | Published:

Protein interaction screening by quantitative immunoprecipitation combined with knockdown (QUICK)

Nature Methods volume 3, pages 981983 (2006) | Download Citation

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Abstract

Present screening methods for protein-protein interactions (PPIs) rely on the overexpression of artificial fusion proteins, making it difficult to assess in vivo relevance. Here we combine stable isotope labeling with amino acids in cell culture (SILAC), RNA interference (RNAi), coimmunoprecipitation and quantitative mass-spectrometry analysis to detect cellular interaction partners of endogenous proteins in mammalian cells with very high confidence. We used this screen to identify interaction partners of β-catenin and Cbl.

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Acknowledgements

We thank B. Blagoev from the Center of Experimental Bioinformatics for fruitful discussions, J.V. Olsen and other members of our department for discussion and assistance, H. Clevers for generously providing us with the inducible β-catenin knockdown cell line and the Interaction Proteome project of the European Union for funding.

Author information

Affiliations

  1. Max Planck Institute of Biochemistry, Department of Proteomics and Signal Transduction, Am Klopferspitz 18, D-82152 Martinsried, Germany.

    • Matthias Selbach
    •  & Matthias Mann

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Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Matthias Mann.

Supplementary information

Word documents

  1. 1.

    Supplementary Table 1

    Sequences and normalized abundance ratios of the quantified peptides for β-catenin and three detected interaction partners.

  2. 2.

    Supplementary Table 2

    Sequences and normalized abundance ratios of the quantified peptides for Cbl and four detected interaction partners.

  3. 3.

    Supplementary Table 3

    Additional proteins identified with increased abundance ratios in the Cbl pulldown.

  4. 4.

    Supplementary Table 4

    Ratio of proteins identified in the whole cell lysates after Cbl knock-down.

  5. 5.

    Supplementary Methods

About this article

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DOI

https://doi.org/10.1038/nmeth972

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