Unraveling Kinase Activation Dynamics Using Kinase-Substrate Relationships from Temporal Large-Scale Phosphoproteomics Studies
Fig 4
Validation of IRS1 S265 as an AKT substrate.
A) Comparison of AKT and RPS6KB1 consensus motif and IRS1 S265 site. B) CTA of AKT (green) and RPS6KB1 (purple) and time profile of IRS1 S265 (blue). (CTA is depicted with mean ± SD) C) Scatter plot of RPS6KB1 prediction scores (y-axis) against RPS6KB1 prediction score—AKT prediction score (x-axis). AKT training substrates are shown in red and RPS6KB1 training substrates are shown in blue. IRS1 S265 is shown in green. D) Insulin signaling via AKT and RPS6KB1. See main text for details. E) 3T3-L1 adipocytes were stimulated with insulin alone or in the presence of inhibitors of AKT (MK, GDC) or mTORC1 (Rapa), after which AKT and RPS6KB1 signaling were assessed by Western blotting. Blots shown are representative of 3 separate experiments. F) Quantification of IRS1 S265 phosphorylation from (E), depicted as mean ± SEM.