Rapamycin inhibits mSin1 phosphorylation independently of mTORC1 and mTORC2.
Current knowledge indicates that the mammalian target of rapamycin (mTOR) functions as two complexes, mTORC1 and mTORC2, regulating cell growth, proliferation, survival, differentiation, and motility. Recently mSin1 has been identified as a critical component of mTORC2, which is essential for phosphorylation of Akt and other signaling molecules. Studies have shown that rapamycin inhibits phosphorylation of mSin1. However, the underlying mechanism is unknown. Here we found that rapamycin inhibited phosphorylation of mSin1 potently and rapidly. Expression of rapamycin-resistant mutant of mTOR (mTOR-T), but not rapamycin-resistant and kinase dead mutant of mTOR (mTOR-TE), prevented rapamycin from inhibiting mSin1 phosphorylation, suggesting that rapamycin-induced dephosphorylation of mSin1 is mTOR-dependent. Surprisingly, ectopic expression of rapamycin-resistant and constitutively active p70 S6 kinase 1 (S6K1) did not confer resistance to rapamycin-induced dephosphorylation of mSin1. Furthermore, disruption of mTORC1 and mTORC2 by silencing raptor and rictor, respectively, or downregulation of S6K1 or Akt did not induce the dephosphorylation of mSin1 as rapamycin did. However, silencing mTOR or mLST8 mimicked the effect of rapamycin, inhibiting mSin1 phosphorylation. Our findings suggest that rapamycin inhibits mSin1 phosphorylation, which is independent of mTORC1 and mTORC2, but is possibly dependent on a new mTOR complex, which at least contains mTOR and mLST8.
State Key Laboratory of Biotherapy/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, People's Republic of China.|Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, Shreveport, LA, USA.|Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, LA, USA.
Yan
Y
Luo
Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, Shreveport, LA, USA.
Lei
L
Liu
State Key Laboratory of Biotherapy/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, People's Republic of China.|Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, Shreveport, LA, USA.
Yang
Y
Wu
Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, Shreveport, LA, USA.
Karnika
K
Singh
Department of Immunobiology, Yale University School of Medicine, New Haven, CT, USA.
Bing
B
Su
State Key Laboratory of Biotherapy/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, People's Republic of China.
Nan
N
Zhang
State Key Laboratory of Biotherapy/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, People's Republic of China.
Xiaowei
X
Liu
State Key Laboratory of Biotherapy/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, People's Republic of China.
Yangmei
Y
Shen
Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, Shreveport, LA, USA.|Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, LA, USA.
Shile
S
Huang
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D006801:Humans; D000076222:Mechanistic Target of Rapamycin Complex 1; D000076225:Mechanistic Target of Rapamycin Complex 2; D020559:Monomeric GTP-Binding Proteins; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D034741:RNA, Small Interfering; D020123:Sirolimus / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D014162:Transfection
D000903:Antibiotics, Antineoplastic; D002352:Carrier Proteins; D046912:Multiprotein Complexes; D034741:RNA, Small Interfering; C000603431:mSin1 protein, human; D000076222:Mechanistic Target of Rapamycin Complex 1; D000076225:Mechanistic Target of Rapamycin Complex 2; D058570:TOR Serine-Threonine Kinases; D020559:Monomeric GTP-Binding Proteins; D020123:Sirolimus