Protein Proximity Mapping: Getting to Know Your Neighbors.

Compartmentalization is essential for all complex forms of life. In eukaryotic cells, membrane-bound organelles, as well as a multitude of protein- and nucleic acid-rich subcellular structures, maintain boundaries and serve as enrichment zones to promote and regulate protein function, including signaling events. Understanding the composition of each cellular "compartment" (be it a classical organelle or a large protein complex) remains a challenging task. For soluble protein complexes, different approaches coupled to mass spectrometry such as affinity purification, biochemical fractionation and proximity labeling of proteins in vivo using biotin-transfer provide important insight. For detergent-insoluble components, proximity labeling has been very effective in identifying protein neighbors, leading to rapid uptake of this technique in the scientific community. This presentation will introduce and compare protein-protein interaction mapping techniques and describe how to design, execute, and analyze proximity labeling experiments, with an emphasis on performing these experiments in a core facility. We have generated a human cell map of major organelles and non-membrane-bound structures from proteins profiled by in vivo biotinylation (BioID). This resource is now a community tool, and I will discuss how it can be used to better interpret BioID results. Since its introduction less than a decade ago, there has been tremendous progress in applying and improving proximity labeling techniques, and I will conclude with a glimpse of what is next on the horizon for proximity mapping.