AMPK-TBC1D1 Nxus Regulates The Secretion Of IGF1 And The Progress Of Metabolic Associated Diseases

With the development of global economy and the improvement of people’s living standard,more people in modern society appear in varying degrees of metabolic syndrome symptoms which seriously affect people’s health and quality of life.AMPK(AMP-activated protein kinase)is a very important protein kinase which can sense energy status and then regulate catabolism in mammalians.The change on the acivity of AMPK will regulate the progress of metabolic syndrome directly or indirectly.TBC1D1(Tre-2/USP6,BUB2,Cdc16 domain family member 1)is a small G protein activating protein and previous study indicates that AMPK can regulate the phosphorylation of serine at position 231 of TBC1D1 to alter the GAP enzyme activity of TBC1D1 protein,but so far the physiological role and importance of the regulatory pathway has not been studied clearly.To address this question,we generated a mouse model harboring the whole body mutation of TBC1D1 protein which was inactivating mutation from the serine at position 231 to alanine and shorting for TBC1D1S231A KI mice.The mutant mice can block the phosphorylation of serine at position 231 of TBC1D1 protein by AMPK kinase.We find that the knockin mice develop obesity on a normal chow diet.While older,the knockin mice cause severe metabolic syndrome including hyperglycemia,hypercholesterolemia,hyperinsulinemia,insulin resistance,type 2 diabetes and non-alcohol fatty acid liver disease(NAFLD).Mechanistically,TBC1D1 and Rab8A can be partially located on the IGF1 storage vesicles and then regulate the secretion rate of IGF1.Further studies of TBC1D1S231A KI mice have shown that the mutant mice increase endocrinal and paracrinal/autocrinal IGF1 secretion rate via increasing the proportion of the small G protein Rab8A-GDP binding form.Hypersecretion of IGF1 causes increased synthesis of fatty acid via activating IGF 1 Receptor-PKB(AKT)-TSC2-mTORC1-Lipin1-Srebp1-Lipogenesis signaling pathway in adipose tissue.Also in skeletal muscle,adipose tissue and liver,hypersecretion of IGF 1 protein causes increased synthesis of protein via activating IGF1 Receptor-PKB(AKT)-TSC2-mTORC1-S6K signaling pathway and the other proteins like FOXO1,GSK3 or 4EBP1.Finally,significantly increased systematic growth rate and lipid accumulation in TBC1D1S231A KI mice result in overweight,obesity and associated metabolic syndrome as the knockin mice aging.To further study the regulation of IGF1 secretion by AMPK protein and the related signaling pathways,we find that reducing the activity of LKB1 and AMPK can indirectly increase the secretion of IGF1 and activate the IGF1 Receptor-PKB(AKT)signaling pathway and then increase cellular anabolism in primary hepatocytes.In the meanwhile,in order to further explore the regulatory mechanism of TBC1D1 protein on the secretion of IGF 1,we also constructed a mouse model with whole body deletion of TBC1D1 protein.Through the analysis of TBC1D1 protein knockout mice,we find that due to the knockout of TBC1D1 protein,the mice have a severe lagging growth rate and reduced fatty acid accumulation.But until now the specific mechanism that causes the phenotype is still in the study.In order to further verify that the obesity in TBC1D1S231A KI mice is caused by increased secretion of IGF1,We constructed a TBC1D1S231A/IGF1+/-mouse model which was knocked out one allele of IGF 1 gene on the basis of TBC1D1S231A KI mice to restore the systemic IGF 1 secretion level to wild-type mice.The results show that TBC1D1S23A/IGF1+/-mice can significantly inactivate IGF1 Receptor-PKB(AKT)-TSC2-mTORC1-Lipin1-Srebp1-Lipogenesis signaling pathway by restoring the secretion level of IGF1 and ultimately reduce fatty acid accumulation in adipose tissue.In addition,the protein synthesis and growth rate of bone in the mice can also be mitigated by inactivating the IGF1 Receptor-PKB(AKT)-TSC2-mTORC1-S6K signaling pathway.Eventually,TBC1D1S231A/IGF1+/-mice have a phenotype with slow growth rate and resistance to obesity relative to TBC1D1S231A KI mice.Based on all the above findings,by constructing and analyzing TBC1D1S231A KI mice and TBC1D1S231A/IGF1+/-mice,we find that systemic blocking the activity of the AMPK-TBC1D1 signaling pathway will up regulate the endocrinal and paracrinal/autocrinal IGF1 secretion rate via increasing the proportion of Rab8A-GDP binding form.Hypersecretion of IGF1 significantly increases the synthesis of fatty acid and protein by activating IGF1 Receptor-PKB(AKT)-TSC2-mTORC1-related signaling pathways.With age,mice will appear in the metabolic syndrome including hypercholesterolemia,hyperglycemia,hyperinsulinemia,insulin resistance and non-alcoholic fatty liver disease and so on.Collectively,in this study,the regulation of IGF1 secretion via AMPK-TBC1D1-Rab8A signaling pathway is first proposed by using the related mouse models.These findings greatly improve the cellular functions of LKB1,AMPK and Rab8A.The study effectively answers the physiological significance of the phosphorylation event on TBC1D1 protein by AMPK,which is to balance the systematic homeostasis of glucose and fatty acid metabolism by regulating the secretion of IGF1.The results also analyze new mechanism on the secretion of IGF 1,which provides new ideas and methods for studying the physiological function of IGF1.The research suggests that the organism can balance the homeostasis between the LKB1-AMPK-TBC1D1 signaling pathway and the IGF1 Receptor-PKB(AKT)-mTORC1 signaling pathway by adjusting the secretion level of IGF1.In mammalians,the abnormalities of the regulatory mechanism will change the body’s synthesis of fatty acid and protein,and then regulate the process of obesity and metabolic syndrome in the case of normal diet.The clarification of the mechanism provides new ideas and methods for the future development of related drugs and treatment of related diseases.