Skeletal muscle insulin resistance is the earliest defect in type 2 diabetes (T2D), however, whether this represents a cell-autonomous defect or is secondary to changes in hormones and other circulating factors in vivo is unclear. To identify primary drivers of skeletal muscle insulin resistance in T2D without interference of these systemic factors, we have developed a “disease-in-a-dish” model by differentiating induced pluripotent stem cells (iPSCs) from T2D patients and controls into skeletal myoblasts (iMyos) and studied their function in vitro (n=8 subjects per group). We find that T2D iMyos retain multiple features of in vivo insulin resistance including altered insulin signaling downstream of the IRS/AKT pathway, impaired insulin-stimulated glucose uptake, and reduced mitochondrial oxidation. More importantly, using global phosphoproteomics we identify nearly 1,000 phosphorylation sites that were significantly increased or decreased (FDR < 0.05) by T2D status. While many of the observed signaling defects in T2D iMyos occurred inside the classical insulin signaling pathway and included dysregulation of Akt, mTOR and forkhead box transcription factors, over 85% of these changes occurred in basal protein phosphorylation. These included phosphorylation of proteins in many pathways not typically involved in insulin resistance, such as mRNA splicing and processing, gene transcription, chromatin remodeling and cytoskeleton dynamics. Motif analysis revealed enrichment of a multiplicity of kinases including ROCK, mTOR/S6K and PKCs with potential targets involved in insulin action, cytoskeleton and cell cycle, without changes in total protein content. This broad dysregulated phosphorylation network reveals a new dimension in the complexity of the cell-autonomous mechanisms underlying insulin resistance in T2D, as well as new targets for therapy or prevention of the disease.


T.M. Batista: None. N.J. Wewer Albrechtsen: Research Support; Self; Mercodia, Novo Nordisk A/S, Novo Nordisk Foundation. Speaker’s Bureau; Self; Merck Sharp & Dohme Corp. J.R. Zierath: None. M. Mann: None. C. Kahn: Advisory Panel; Self; ERX Pharmaceuticals, Kaleido Biosciences. Consultant; Self; AntriaBio, Flagship Pioneering, Sana-Cobalt.


National Institutes of Health (R01DK031036, R01DK033201); Swedish Research Council (2015-00165); Novo Nordisk Foundation (NNF18CC0034900)

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