Role of mTORC1 in lysosomal localization in glucagon secretion
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BACKGROUND: Elevation of glucagon levels and increase in alpha-cell mass are associated with states of hyperglycemia in diabetes. However, little is known about the mechanisms that control glucagon secretion and alpha-cell mass expansion in normal or diabetogenic conditions. Glucagon is secreted during the fasting state, when glucose levels are low, to stimulate glycogenolysis and gluconeogenesis in the liver to increase the blood glucose level. Amino acids, also, stimulate-glucagon secretion and alpha-cell mass. Amino acids increase glucagon secretion via activation of mTORC1 in alpha-cells. A critical step for mTOR activation is the localization of mTORC1 to the lysosome where it meets Rheb for activation. Amino acids are unique in their ability to localize mTORC1 to the lysosomal membrane for activation through their interaction with a variety of amino acid sensors, such as Sestrin2, which modulates mTORC1 activity via its interaction with GATOR2. Integral to mTORC1’s localization is the Ragulator complex, more specifically, p18, which provides the essential scaffolding necessary for lysosomal docking. Amino acids sensors work upstream of mTORC1 and sense amino acid concentrations with different affinities and are specific to certain amino acids and relay this information to mTORC1. OBJECTIVE: To investigate the role that p18, a component of the Ragulator complex, and GATOR2, a component of an amino acid sensor complex, play in amino-acid dependent mTORC1 lysosomal localization and its effect on alpha-cell function and glucagon secretion. METHODS: Generation of Knockout mice for p18 and GATOR2 in alpha-cells were produced by crossing Glu-Cre mice with P18(flox/flox) and GATOR2(flox/flox). Blood glucose, glucagon, and insulin levels were evaluated during fed, fasting, and insulin-induced hypoglycemic conditions to evaluate glucagon secretion. Isolated islets were also exposed to media containing different glucose or nutrient concentrations to evaluate the effect on glucagon secretion. RESULTS: Our data shows that knockdown experiments in alpha-cells for p18 and GATOR2 have demonstrated the role of these proteins in amino acid dependent localization of mTORC1 to the lysosomal membrane. More specifically, our data demonstrate that animals with a knockdown for p18 or GATOR2 demonstrated decreased glucagon secretion during hypoglycemic conditions. Mice with a knockdown for p18 also demonstrate decreased glucagon secretion in the presence of glucagon secretion stimulators such as arginine and also demonstrated decreased insulin secretion. CONCLUSIONS: Loss of essential components of the amino acid signaling and lysosomal localization in the mTORC1 pathway results in impaired function of alpha-cells and glucagon secretion. Loss of p18 in alpha-cells potentially results in an inability of mTORC1 to dock and bind to the lysosomal membrane, whereas loss of GATOR2 potentially results in chronic inhibition of mTORC1 via GATOR1. Loss of each of these components results in the lost or impaired ability for mTORC1 to migrate and bind to the lysosomal membrane.