SUN-593 - A Kinase Mediated Signaling and Glucose Uptake Defect in Human Insulin Resistance

Insulin resistance is a major risk factor in the development of type 2 diabetes (T2D) and metabolic syndrome. Although ~25% of people within the general non-diabetic population are insulin resistant, the primary underlying cause of insulin resistance remains elusive. In this study, we have used induced pluripotent stem cells (iPSC) derived from non-diabetic humans at both ends of the insulin sensitivity spectrum, i.e., the top 20% of insulin resistance vs. the top 20% of insulin sensitivity differentiated into myoblasts (iMyos) to model insulin resistance in vitro. Global phosphoproteomics analysis of these cells showed a large network of dysregulated protein phosphorylations linked to differences in insulin sensitivity including 378 up-regulated and 393 down-regulated insulin stimulated phosphosites in I-Res iMyos (p < 0.05). To identify drivers of altered phosphorylation in insulin resistance, we used an in silico AI kinome analysis to query which of the 300+ Ser/Thr kinases encoded in the human genome might be responsible for the protein phosphorylation changes. We could identify 19 kinases whose predicted activities were significantly increased in I-Res iMyos, suggesting their potential link to the pathogenesis of insulin resistance. To functionally link these altered kinases to the downstream defect of impaired glucose uptake associated with insulin resistance, we conducted a loss-of-function screen using a CRISPR-based approach to identify candidate kinases, phosphatases or their regulators that positively or negatively regulated glucose uptake in I-Res iMyos. Among the 19 kinases predicted to have increased activity in insulin resistance, one kinase, the Dual Specificity Tyrosine Phosphorylation Regulated Kinase 2 (DYRK2), was identified as a potential modulator of insulin resistance by being both increased in predicted activity in I-Res iMyos and associated with rescued glucose uptake in I-Res iMyos by CRISPR-mediated knockdown. We found important DYRK2 substrates involved in the glucose transport mechanism and a reciprocal relationship between DYRK2 and Glut1 and Glut 4 protein levels in I-Res iMyos. In summary, combining a Kinome analysis with CRISPR screening reveals DYRK2 as an important upstream regulator of human insulin resistance.

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