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Marked by association: techniques for proximity-dependent labeling of proteins in eukaryotic cells

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Abstract

Various methods have been established for the purpose of identifying and characterizing protein–protein interactions (PPIs). This diverse toolbox provides researchers with options to overcome challenges specific to the nature of the proteins under investigation. Among these techniques is a category based on proximity-dependent labeling of proteins in living cells. These can be further partitioned into either hypothesis-based or unbiased screening methods, each with its own advantages and limitations. Approaches in which proteins of interest are fused to either modifying enzymes or receptor sequences allow for hypothesis-based testing of protein proximity. Protein crosslinking and BioID (proximity-dependent biotin identification) permit unbiased screening of protein proximity for a protein of interest. Here, we evaluate these approaches and their applications in living eukaryotic cells.

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Abbreviations

BAT:

Biotin acceptor tag

BioID:

Proximity-dependent biotin identification

BLINC:

Biotin labeling of intercellular contacts

ChIP:

Chromatin immuno-precipitation

FRET:

Forster (fluorescence) resonance energy transfer

ID-PRIME:

Interaction-dependent probe incorporation mediated by enzymes

LAP:

LplA acceptor peptide

PIR:

Protein interaction reporter

PPI:

Protein–protein interaction

PUB-MS:

Proximity utilizing biotinylation and mass spectrometry

PUB-NChIP:

Proximity utilizing biotinylation with native ChIP

TRAP:

Targeted releasable affinity probe

Y2H:

Yeast-2-hybrid

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Acknowledgment

This work was supported by Sanford Research.

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Authors and Affiliations

  1. Children’s Health Research Center, Sanford Research/USD, North 60th St. East, Sioux Falls, SD, 57104, USA

    Kyle J. Roux

  2. Department of Pediatrics, University of South Dakota School of Medicine, Sioux Falls, SD, 57105, USA

    Kyle J. Roux

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Correspondence to Kyle J. Roux.

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Roux, K.J. Marked by association: techniques for proximity-dependent labeling of proteins in eukaryotic cells. Cell. Mol. Life Sci. 70, 3657–3664 (2013). https://doi.org/10.1007/s00018-013-1287-3

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