Studies Of Systemic Growth Regulation By Tre2-Bud2-Cdc16 (TBC) Domain Family,Member 7 (TBC1D7)

The insulin pathway integrates the nutrition viability to control systemic growth in multiple organism development.Meanwhile insulin pathway affects the fitness and lifespan by regulating stress resistance and tissue homeostasis.TSC complex(Tuberous Sclerosis Complex)likes to be the central node between insulin signaling pathway and TOR pathway which regulates cell growth.TSC complex contains TSC1(Tuberous Sclerosis protein 1),TSC2(Tuberous Sclerosis protein 2)and TBC1D7(Tre2-Bud2-Cdc16(TBC)domain family,member 7).It is known that TBC1D7 binds to TSC1,however,the physiological functions of TBC1D7 in vivo are still lacking.dTBC1D7(coded by CG6182 in Drosophila)was a close homolog of human TBC1D7.A null mutant fly of dTBC1D7,named as dTBC1D7KO has been generated by using CRISPR/Cas9.dTBC1D7KO flies exhibited larger body sizes than wild-type flies at both the pupa and adult stages.The overgrowth phenotypes were completely rescued by the ubiquitous expression of dTBC1D7.The overgrowth was a type of systemic growth in a cell-non-autonomous manner.The mRNA level of ilp2(Insulin like peptide 2)was significantly up-regulated in dTBC1D7KO flies,and the expression pattern of ILP2 in the dTBC1D7KO was changed,indicating that ILP2 secretion was stimulated.Further studies suggested that the level of circulatory ILP2 was significantly increased in mutant flies and the level of circulatory glucose was decreased.Taken together,our data indicated that the circulatory insulin was elevated in the dTBC1D7 mutant flies.ILPs are synthesized and secreted in the IPCs(insulin producing cells)in the brain of Drosophila.dTBClD7 was mainly expressed in the brain and ovary of flies and it has little expression in other tissues.Through staining the brain of flies,dTBC1D7 was largely enriched in the IPC neurons.Encouragingly,expression of dTBC1D7 and mammal TBC1D7 in IPC neurons was able to rescue the systemic overgrowth of dTBC1D7KO;moreover,genetic epstasis analysis indicated that the systemic growth promotion caused by dTBC1D7 mutation was dependent on ilp2.Meanwhile,dTBC1D7I125Q(a dTBC1D7 mutant isoform discarded binding activity to TSC1)also rescued the overgrowth phenotype to the same extent as wild-type dTBC1D7 did.Therefore,dTBC1D7 functioned in IPC neurons to regulate growth through ILP2 and this function was independent on TSC complex.Finally,dTBC1D7 mutation animals were short-lived and more sensitive to stress by increasing the insulin signaling transduction.TBC1D7 knockout mouse was generated and the TBC1D7KO mouse also exhibited systemic growth promotion compared to wild-type.This phenotpye implied that the machanisms of TBC1D7 in growth control would be conserved between fly and mammals.This presented work provide new insights of the physiological function of TBC1D7 in the systemic control of growth,metabolism,life span,and stress resistance by controlling ILP2 biogenesis.This work also provide some interesting implications in the human disorders caused by TBC1D7 mutation.