High-throughput phosphoproteomics reveals in vivo insulin signaling dynamics

Nat Biotechnol. 2015 Sep;33(9):990-5. doi: 10.1038/nbt.3327. Epub 2015 Aug 17.


Mass spectrometry has enabled the study of cellular signaling on a systems-wide scale, through the quantification of post-translational modifications, such as protein phosphorylation. Here we describe EasyPhos, a scalable phosphoproteomics platform that now allows rapid quantification of hundreds of phosphoproteomes in diverse cells and tissues at a depth of >10,000 sites. We apply this technology to generate time-resolved maps of insulin signaling in the mouse liver. Our results reveal that insulin affects ~10% of the liver phosphoproteome and that many known functional phosphorylation sites, and an even larger number of unknown sites, are modified at very early time points (<15 s after insulin delivery). Our kinetic data suggest that the flow of signaling information from the cell surface to the nucleus can occur on very rapid timescales of less than 1 min in vivo. EasyPhos facilitates high-throughput phosphoproteomics studies, which should improve our understanding of dynamic cell signaling networks and how they are regulated and dysregulated in disease.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cells, Cultured
  • Gene Expression Profiling / methods
  • High-Throughput Screening Assays / methods*
  • Insulin / metabolism*
  • Liver / metabolism*
  • Male
  • Mass Spectrometry / methods
  • Mice
  • Mice, Inbred C57BL
  • Phosphoproteins / metabolism*
  • Protein Interaction Mapping / methods
  • Proteome / metabolism*
  • Signal Transduction / physiology*


  • Insulin
  • Phosphoproteins
  • Proteome