Cytoplasmic and nuclear distribution of the protein complexes mTORC1 and mTORC2: rapamycin triggers dephosphorylation and delocalization of the mTORC2 components rictor and sin1.
The mammalian target of rapamycin (mTOR) is part of two distinct complexes, mTORC1, containing raptor and mLST8, and mTORC2, containing rictor, mLST8 and sin1. Although great endeavors have already been made to elucidate the function and regulation of mTOR, the cytoplasmic nuclear distribution of the mTOR complexes is unknown. Upon establishment of the proper experimental conditions, we found mTOR, mLST8, rictor and sin1 to be less abundant in the nucleus than in the cytoplasm of non-transformed, non-immortalized, diploid human primary fibroblasts. Although raptor is also high abundant in the nucleus, the mTOR/raptor complex is predominantly cytoplasmic, whereas the mTOR/rictor complex is abundant in both compartments. Rapamycin negatively regulates the formation of both mTOR complexes, but the molecular mechanism of its effects on mTORC2 remained elusive. We describe that in primary cells short-term treatment with rapamycin triggers dephosphorylation of rictor and sin1 exclusively in the cytoplasm, but does not affect mTORC2 assembly. Prolonged drug treatment leads to complete dephosphorylation and cytoplasmic translocation of nuclear rictor and sin1 accompanied by inhibition of mTORC2 assembly. The distinct cytoplasmic and nuclear upstream and downstream effectors of mTOR are involved in many cancers and human genetic diseases, such as tuberous sclerosis, Peutz-Jeghers syndrome, von Hippel-Lindau disease, neurofibromatosis type 1, polycystic kidney disease, Alzheimer's disease, cardiac hypertrophy, obesity and diabetes. Accordingly, analogs of rapamycin are currently tested in many different clinical trials. Our data allow new insights into the molecular consequences of mTOR dysregulation under pathophysiological conditions and should help to optimize rapamycin treatment of human diseases.
Rosner
Margit
M
Medical Genetics, Medical University of Vienna, Währinger Gürtel 18-20, 1090 Vienna, Austria.
Hengstschläger
Markus
M
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D000076226:Rapamycin-Insensitive Companion of mTOR Protein; D020123:Sirolimus / Q000494:pharmacology*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C500984:CRTC1 protein, human; C493245:CRTC2 protein, human; D002352:Carrier Proteins; C493277:MAPKAP1 protein, human; C492571:RICTOR protein, human; D000076226:Rapamycin-Insensitive Companion of mTOR Protein; D014157:Transcription Factors; D020123:Sirolimus