Bo Yu1, Pei Xu1, Zhenyang Zhao1, Jiyang Cai1, Paul Sternberg2, Yan Chen1. 1. Department of Ophthalmology and Visual Sciences, University of Texas Medical Branch, Galveston, Texas, United States. 2. Vanderbilt Eye Institute, Vanderbilt University, Nashville, Tennessee, United States.
Abstract
PURPOSE: Inhibiting mechanistic target of rapamycin (mTOR) by pharmacological or genetic approaches can extend lifespan in mammals. The kinase activity of mTOR is controlled by upstream regulatory proteins and its subcellular localization. The purpose of this study was to characterize age-related alterations and functional consequences of mTOR signaling in the postmitotic RPE cells. METHODS: Activity of mTOR complex 1 (mTORC1) was monitored by measuring phosphorylation status of its downstream effector protein S6, in either cultured human RPE cells or RPE explants prepared from mice at different ages. Subcellular distribution of mTOR was investigated by immunofluorescent staining of RPE culture or flatmount. The signaling of mTORC1 was modulated by either overexpression of a small guanosine triphosphatase, Ras homolog enriched in brain (Rheb), or disruption of the Ragulator complex with small interference RNA targeting p18. The effects of mTOR pathway on degradation of phagocytosed photoreceptor outer segments (POS) were determined by measuring the turnover rate of rhodopsin. RESULTS: Aged RPE cells had more lysosome-associated mTOR and had increased response to amino acid stimulation. The lysosome distribution was essential for mTORC1 function, as disruption of the Ragulator complex abolished mTORC1 activation by amino acids. Increased mTORC1 activity caused decreased rate of degradation of internalized POS in the RPE. CONCLUSIONS: Aging changes the subcellular localization and function of mTOR in the RPE. Increased mTORC1 inhibits POS degradation and may further exacerbate lysosome dysfunction of aged RPE. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.
PURPOSE: Inhibiting mechanistic target of rapamycin (mTOR) by pharmacological or genetic approaches can extend lifespan in mammals. The kinase activity of mTOR is controlled by upstream regulatory proteins and its subcellular localization. The purpose of this study was to characterize age-related alterations and functional consequences of mTOR signaling in the postmitotic RPE cells. METHODS: Activity of mTOR complex 1 (mTORC1) was monitored by measuring phosphorylation status of its downstream effector protein S6, in either cultured human RPE cells or RPE explants prepared from mice at different ages. Subcellular distribution of mTOR was investigated by immunofluorescent staining of RPE culture or flatmount. The signaling of mTORC1 was modulated by either overexpression of a small guanosine triphosphatase, Ras homolog enriched in brain (Rheb), or disruption of the Ragulator complex with small interference RNA targeting p18. The effects of mTOR pathway on degradation of phagocytosed photoreceptor outer segments (POS) were determined by measuring the turnover rate of rhodopsin. RESULTS: Aged RPE cells had more lysosome-associated mTOR and had increased response to amino acid stimulation. The lysosome distribution was essential for mTORC1 function, as disruption of the Ragulator complex abolished mTORC1 activation by amino acids. Increased mTORC1 activity caused decreased rate of degradation of internalized POS in the RPE. CONCLUSIONS: Aging changes the subcellular localization and function of mTOR in the RPE. Increased mTORC1 inhibits POS degradation and may further exacerbate lysosome dysfunction of aged RPE. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.
Authors: John E Wilkinson; Lisa Burmeister; Susan V Brooks; Chi-Chao Chan; Sabrina Friedline; David E Harrison; James F Hejtmancik; Nancy Nadon; Randy Strong; Lauren K Wood; Maria A Woodward; Richard A Miller Journal: Aging Cell Date: 2012-06-04 Impact factor: 9.304
Authors: Tamir Chandra; Kristina Kirschner; Jean-Yves Thuret; Benjamin D Pope; Tyrone Ryba; Scott Newman; Kashif Ahmed; Shamith A Samarajiwa; Rafik Salama; Thomas Carroll; Rory Stark; Rekin's Janky; Masako Narita; Lixiang Xue; Agustin Chicas; Sabrina Nũnez; Ralf Janknecht; Yoko Hayashi-Takanaka; Michael D Wilson; Aileen Marshall; Duncan T Odom; M Madan Babu; David P Bazett-Jones; Simon Tavaré; Paul A W Edwards; Scott W Lowe; Hiroshi Kimura; David M Gilbert; Masashi Narita Journal: Mol Cell Date: 2012-07-12 Impact factor: 17.970
Authors: Oliver Dreesen; Alexandre Chojnowski; Peh Fern Ong; Tian Yun Zhao; John E Common; Declan Lunny; E Birgitte Lane; Shu Jin Lee; Leah A Vardy; Colin L Stewart; Alan Colman Journal: J Cell Biol Date: 2013-02-25 Impact factor: 10.539
Authors: Akintomide Apara; Joana Galvao; Yan Wang; Murray Blackmore; Allison Trillo; Keiichiro Iwao; Dale P Brown; Kimberly A Fernandes; Abigail Huang; Tu Nguyen; Masoumeh Ashouri; Xiong Zhang; Peter X Shaw; Noelia J Kunzevitzky; Darcie L Moore; Richard T Libby; Jeffrey L Goldberg Journal: J Neurosci Date: 2017-09-04 Impact factor: 6.167
Authors: Cheng-Kang Chiang; Aleksander Tworak; Brian M Kevany; Bo Xu; Janice Mayne; Zhibin Ning; Daniel Figeys; Krzysztof Palczewski Journal: J Biol Chem Date: 2017-10-04 Impact factor: 5.157
Authors: Yan Wang; Evan G Cameron; Jinliang Li; Travis L Stiles; Michael D Kritzer; Rahul Lodhavia; Jonathan Hertz; Tu Nguyen; Michael S Kapiloff; Jeffrey L Goldberg Journal: EBioMedicine Date: 2015-10-28 Impact factor: 8.143