title: Differentiation of glutamatergic neurons from mouse embryonic stem cells requires raptor S6K signaling.
abstract: Although the mammalian target of rapamycin complex 1 (mTORC1) functions as an important signaling complex in many cellular processes, the role of mTORC1 in neurons derived from embryonic stem cells (ESCs) has been less explored. Here, using a modified protocol to differentiate mouse ESCs (mESCs) into almost uniform glutamatergic neurons, we explored the importance of raptor/mTORC1 in the differentiation of mESCs. Raptor gene-trap mESCs, and raptor-knockdown mESCs formed smaller-sized embryonic bodies than the wild type and failed to undergo neuronal differentiation. Treatment with 1μM rapamycin starting at the point when neuronal precursors began to differentiate from mESCs caused the gradual loss of neurites, shrinkage of soma, and a decreased ratio of neurite length to cell number over 48 to 72h of treatment. This change was accompanied by activation of caspase-3 and S6 kinase (S6K), but not 4E-binding protein 1 (4EBP1). Knockdown of raptor during neuronal differentiation from mESCs also resulted in gradual loss of neurites and shrinkage of cell bodies. Loss of neurite density resulting from rapamycin treatment could be reversed by overexpression of S6K T389E. Taken together, these data demonstrate that raptor/mTORC1/S6K plays a critical role in the differentiation and survival of neurons derived from mESCs.
authors.affiliation: Department of Life Science, National Taiwan Normal University, Taipei 116,Taiwan.
authors.forename: Jen-Hua
authors.initials: JH
authors.lastname: Chuang
authors.forename: Li-Chu
authors.initials: LC
authors.lastname: Tung
authors.forename: Yi
authors.initials: Y
authors.lastname: Yin
authors.forename: Yenshou
authors.initials: Y
authors.lastname: Lin
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D018797:Cell Cycle Proteins; D002454:Cell Differentiation / Q000187:drug effects; D048429:Cell Size; D058732:Embryoid Bodies / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D039642:Eukaryotic Initiation Factors; D007166:Immunosuppressive Agents / Q000494:pharmacology; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D008807:Mice, Inbred BALB C; D046912:Multiprotein Complexes / Q000378:metabolism; D009154:Mutation; D016501:Neurites / Q000378:metabolism; D009474:Neurons / Q000166:cytology* / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D000076223:Regulatory-Associated Protein of mTOR; D019893:Ribosomal Protein S6 Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; C499527:Eif4ebp1 protein, mouse; D039642:Eukaryotic Initiation Factors; D007166:Immunosuppressive Agents; D046912:Multiprotein Complexes; D010750:Phosphoproteins; D034741:RNA, Small Interfering; D000076223:Regulatory-Associated Protein of mTOR; C540162:Rptor protein, mouse; D000076222:Mechanistic Target of Rapamycin Complex 1; D019893:Ribosomal Protein S6 Kinases; D058570:TOR Serine-Threonine Kinases; D053148:Caspase 3; D020123:Sirolimus