Additional file 14: of Glucose-dependent phosphorylation signaling pathways and crosstalk to mitochondrial respiration in insulin secreting cells

Figure S6. KSEA output robustness. Groups of p-sites defining the regulation of specific kinases during glucose stimulation are shown in A-F. Changes p-status are plotted as changes over time. The average changes calculated from within these groups reflect the regulation of the corresponding kinases (colored lines). To confirm the signaling events revealed by KSEA, we assessed the phosphorylation status of a number of kinase substrates using phospho specific antibodies (A-F). INS-1E protein lysates were prepared at specific time-points over a time course of 0–60 min of glucose activation. The abundance of the total protein was not affected by glucose over the time window studied here as demonstrated using antibodies recognizing the unmodified proteins. For all phospho-specific antibodies tested, Western blotting analysis and quantitative phospho-proteomics agree regarding the changes in phosphorylation status following glucose stimulation. (PDF 485 kb