Insulin sensitivity in skeletal muscle

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Project Summary/Abstract Diabetes, a condition characterized by abnormally low insulin-stimulated glucose transport into skeletal muscle, is a major threat to health. Activation of AMPK, such as by exercise, muscle contractions, or serum starvation, provides a means to increase sensitivity of glucose transport to stimulation by insulin. While enhanced phosphorylation of AS160 occurs concomitant with development of insulin sensitivity, the complete molecular mechanisms underlying the increase in insulin-stimulated glucose transport remain unknown. We aim to fill this important knowledge gap by testing the hypothesis that mTOR and ULK1 mediate AMPK- induced insulin sensitivity. Our preliminary data show that inhibition of mTOR complex 1 (mTORC1) causes an increase in insulin-stimulated glucose transport. We have also found that inhibition of ULK1 prevents insulin- stimulated glucose uptake by serum-starved myotubes, while a pharmacological activator of ULK1 increases AS160 phosphorylation. Finally, our data show that activation of AMPK causes an increase in mTORC2 activity in myotubes, as demonstrated by increased phosphorylation of the mTORC2 substrate Akt. To test our hypotheses, we propose three specific aims. For Aim 1, to determine the role of mTORC1 in insulin sensitivity, we will test the hypothesis that inhibition of mTORC1 causes increased insulin sensitivity and mediates AMPK- induced insulin sensitivity. Approaches for this aim will include determining whether inhibition of mTORC1 by pharmacological and genetic means will increase insulin-stimulated glucose transport and whether activation of mTORC1 by physiological and genetic means will prevent development of insulin sensitivity after activation of AMPK, serum starvation, or exercise/muscle contractions. For Aim 2, to determine the role of ULK1 in insulin sensitivity, we will test the hypothesis that activation of ULK1 is sufficient to increase insulin sensitivity and necessary to induction of insulin sensitivity by AMPK. Approaches to this aim will include determining whether pharmacological activation of ULK1 increases insulin-stimulated glucose transport and whether pharmacological inhibition of ULK1 or expression of inactive or AMPK-insensitive mutants of ULK1 prevent development of insulin sensitivity. For Aim 3, to determine the role of mTORC2 in AS160 phosphorylation and insulin sensitivity, we will test the hypothesis that mTORC2 is an important mediator of AS160 phosphorylation during development of insulin sensitivity. Approaches will include determining whether pharmacological or genetic disruption of the mTORC2 complex will prevent the normal increase in AS160 phosphorylation after serum starvation, activation of AMPK, or exercise/muscle contractions and whether prior activation of the mTORC2 substrate Akt causes a later increase in insulin-stimulated glucose transport. Completion of the specific aims will elucidate novel roles of mTOR and ULK1 in control of insulin sensitivity in skeletal muscle and suggest future strategies to overcome insulin resistance.