A Cross-Species Study of PI3K Protein-Protein Interactions Reveals the Direct Interaction of P85 and SHP2

Sci Rep. 2016 Feb 3;6:20471. doi: 10.1038/srep20471.

Abstract

Using a series of immunoprecipitation (IP)-tandem mass spectrometry (LC-MS/MS) experiments and reciprocal BLAST, we conducted a fly-human cross-species comparison of the phosphoinositide-3-kinase (PI3K) interactome in a drosophila S2R+ cell line and several NSCLC and human multiple myeloma cell lines to identify conserved interacting proteins to PI3K, a critical signaling regulator of the AKT pathway. Using H929 human cancer cells and drosophila S2R+ cells, our data revealed an unexpected direct binding of Corkscrew, the drosophila ortholog of the non-receptor protein tyrosine phosphatase type II (SHP2) to the Pi3k21B (p60) regulatory subunit of PI3K (p50/p85 human ortholog) but no association with Pi3k92e, the human ortholog of the p110 catalytic subunit. The p85-SHP2 association was validated in human cell lines, and formed a ternary regulatory complex with GRB2-associated-binding protein 2 (GAB2). Validation experiments with knockdown of GAB2 and Far-Western blots proved the direct interaction of SHP2 with p85, independent of adaptor proteins and transfected FLAG-p85 provided evidence that SHP2 binding on p85 occurred on the SH2 domains. A disruption of the SHP2-p85 complex took place after insulin/IGF1 stimulation or imatinib treatment, suggesting that the direct SHP2-p85 interaction was both independent of AKT activation and positively regulates the ERK signaling pathway.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Diptera / metabolism*
  • Drosophila / metabolism*
  • Drosophila Proteins / metabolism
  • Humans
  • Insect Proteins / metabolism
  • Phosphatidylinositol 3-Kinases / metabolism*
  • Protein Binding
  • Protein Tyrosine Phosphatase, Non-Receptor Type 11 / metabolism*
  • Proteomics / methods*
  • Tandem Mass Spectrometry

Substances

  • Drosophila Proteins
  • Insect Proteins
  • Phosphatidylinositol 3-Kinases
  • Protein Tyrosine Phosphatase, Non-Receptor Type 11