Altered mTOR signaling in senescent retinal pigment epithelium.
OBJECTIVE
Mammalian target of rapamycin (mTOR)-mediated pathways play central roles in regulating aging. The purpose of the present study was to characterize the mTOR cascade in human retinal pigment epithelial (RPE) cells and to investigate its potential roles in controlling RPE senescence.


METHODS
Expression of major components of the mTOR signaling networks was evaluated by Western blot analyses. Formations of the two signaling complexes of mTOR, mTORC1, and mTORC2 were determined by coimmunoprecipitation. The activation of mTORC1 was monitored by measuring the phosphorylation status of the downstream substrate protein S6. Senescence of the cultured human RPE cells was assessed by measuring both the senescence associated-β-galactosidase (SA-β-Gal) activity and the expression level of p16, a cyclin-dependent kinase inhibitor.


RESULTS
Human RPE cells contained functional mTORC1 and mTORC2 signaling complexes. The assembly and activity of mTORC1 were regulated by upstream nutrient and growth factor signals. The sensitivity of mTORC1 to extracellular nutrient stimuli increased in RPE cells that had developed in vitro senescence. Suppression of the mTORC1 by rapamycin prevented the appearance of senescence markers in the RPE.


CONCLUSIONS
The mTOR pathway presented age-associated changes in human RPE cells, and downregulation of mTORC1 could delay the aging process of the RPE.
Chen
Yan
Y
Vanderbilt Eye Institute, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA. yan.chen@vanderbilt.edu
Wang
Jian
J
Cai
Jiyang
J
Sternberg
Paul
P
D015153:Blotting, Western; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000502:physiology*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D000076222:Mechanistic Target of Rapamycin Complex 1; D046912:Multiprotein Complexes; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011506:Proteins; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000378:metabolism; D038601:Ribosomal Protein S6 / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D058570:TOR Serine-Threonine Kinases; D014157:Transcription Factors / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
C493245:CRTC2 protein, human; D019941:Cyclin-Dependent Kinase Inhibitor p16; D047908:Intracellular Signaling Peptides and Proteins; D046912:Multiprotein Complexes; D011506:Proteins; D038601:Ribosomal Protein S6; D014157:Transcription Factors; C546842:MTOR protein, human; D000076222:Mechanistic Target of Rapamycin Complex 1; D017346:Protein Serine-Threonine Kinases; D058570:TOR Serine-Threonine Kinases; D001616:beta-Galactosidase
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