title: Inhibition of mTOR Prevents ROS Production Initiated by Ethidium Bromide-Induced Mitochondrial DNA Depletion.
abstract: The regulation of mitochondrial mass and DNA content involves a complex interaction between mitochondrial DNA replication machinery, functional components of the electron transport chain, selective clearance of mitochondria, and nuclear gene expression. In order to gain insight into cellular responses to mitochondrial stress, we treated human diploid fibroblasts with ethidium bromide at concentrations that induced loss of mitochondrial DNA over a period of 7 days. The decrease in mitochondrial DNA was accompanied by a reduction in steady state levels of the mitochondrial DNA binding protein, TFAM, a reduction in several electron transport chain protein levels, increased mitochondrial and total cellular ROS, and activation of p38 MAPK. However, there was an increase in mitochondrial mass and voltage dependent anion channel levels. In addition, mechanistic target of rapamycin (mTOR) activity, as judged by p70S6K targets, was decreased while steady state levels of p62/SQSTM1 and Parkin were increased. Treatment of cells with rapamycin created a situation in which cells were better able to adapt to the mitochondrial dysfunction, resulting in decreased ROS and increased cell viability but did not prevent the reduction in mitochondrial DNA. These effects may be due to a more efficient flux through the electron transport chain, increased autophagy, or enhanced AKT signaling, coupled with a reduced growth rate. Together, the results suggest that mTOR activity is affected by mitochondrial stress, which may be part of the retrograde signal system required for normal mitochondrial homeostasis.
authors.affiliation: Department of Pathology, Drexel University College of Medicine , Philadelphia, PA , USA.
authors.forename: Timothy
authors.initials: T
authors.lastname: Nacarelli
authors.affiliation: Department of Pathology, Drexel University College of Medicine , Philadelphia, PA , USA.
authors.forename: Ashley
authors.initials: A
authors.lastname: Azar
authors.affiliation: Department of Pathology, Drexel University College of Medicine , Philadelphia, PA , USA.
authors.forename: Christian
authors.initials: C
authors.lastname: Sell
keywords: ROS; aging; ethidium bromide; mTOR; mitochondria; p38; rapamycin; senescence
references.citation: Biochem Soc Trans. 2009 Feb;37(Pt 1):217-22
references.pmid: 19143635
references.citation: Science. 2013 Dec 20;342(6165):1524-8
references.pmid: 24231806
references.citation: Nucleic Acids Res. 2002 May 1;30(9):1929-34
references.pmid: 11972329
references.citation: Nature. 2009 Jul 16;460(7253):392-5
references.pmid: 19587680
references.citation: Autophagy. 2012 Apr;8(4):445-544
references.pmid: 22966490
references.citation: Mol Pharmacol. 2007 Jun;71(6):1695-702
references.pmid: 17387142
references.citation: Ann N Y Acad Sci. 2007 Apr;1100:241-5
references.pmid: 17460185
references.citation: Oncol Rep. 2013 Sep;30(3):1201-8
references.pmid: 23828262
references.citation: J Mol Biol. 1969 Aug 28;44(1):195-204
references.pmid: 5811827
references.citation: J Biol Chem. 1983 Aug 10;258(15):9078-85
references.pmid: 6307991
references.citation: Neurotherapeutics. 2013 Apr;10(2):186-98
references.pmid: 23385875
references.citation: Mol Endocrinol. 2000 Jun;14(6):783-94
references.pmid: 10847581
references.citation: Methods Mol Biol. 2009;554:383-91
references.pmid: 19513686
references.citation: Autophagy. 2010 Oct;6(7):871-8
references.pmid: 20798600
references.citation: Nature. 1981 Apr 9;290(5806):457-65
references.pmid: 7219534
references.citation: J Bacteriol. 1975 Aug;123(2):497-504
references.pmid: 1097413
references.citation: Mol Cell Biol. 2009 Aug;29(16):4495-507
references.pmid: 19528227
references.citation: Aging Cell. 2014 Jun;13(3):468-77
references.pmid: 24341993
references.citation: DNA Repair (Amst). 2014 Jul;19:190-8
references.pmid: 24780559
references.citation: J Proteomics. 2009 Aug 20;72(6):1046-60
references.pmid: 19567276
references.citation: Aging Cell. 2013 Dec;12(6):966-77
references.pmid: 23795962
references.citation: PLoS One. 2010 Sep 07;5(9):e12592
references.pmid: 20830296
references.citation: Mitochondrion. 2005 Jun;5(3):173-93
references.pmid: 16050983
references.citation: J Mol Biol. 1969 Nov 28;46(1):201-4
references.pmid: 5358641
references.citation: Toxicol Appl Pharmacol. 2012 May 15;261(1):42-9
references.pmid: 22712077
references.citation: J Biochem. 1998 Jun;123(6):1010-6
references.pmid: 9603986
references.citation: Arch Neurol. 2004 Dec;61(12):1898-904
references.pmid: 15596610
references.citation: J Cell Biol. 1996 Nov;135(4):883-93
references.pmid: 8922374
references.citation: J Biol Chem. 2000 Jul 21;275(29):22558-62
references.pmid: 10811632
references.citation: Mitochondrion. 2009 Jul;9(4):261-5
references.pmid: 19324101
references.citation: PLoS Biol. 2010 Jan 26;8(1):e1000298
references.pmid: 20126261
references.citation: Exp Cell Res. 1972 May;72(1):211-22
references.pmid: 4337144
references.citation: Cell. 1977 Aug;11(4):719-27
references.pmid: 560914
references.citation: Diabetes. 2001 Jan;50(1):24-31
references.pmid: 11147790
references.citation: Mol Cell Biol. 1985 May;5(5):1163-9
references.pmid: 2987677
references.citation: Proc Natl Acad Sci U S A. 1970 Dec;67(4):1926-33
references.pmid: 4321352
references.citation: Nat Biotechnol. 2003 Mar;21(3):281-6
references.pmid: 12592411
references.citation: Mol Genet Metab. 2013 Sep-Oct;110(1-2):35-41
references.pmid: 23911206
references.citation: Exp Cell Res. 2003 Sep 10;289(1):133-42
references.pmid: 12941611
references.citation: Nature. 2004 May 27;429(6990):417-23
references.pmid: 15164064
references.citation: Biochim Biophys Acta. 2010 Feb;1797(2):113-28
references.pmid: 19761752