title: Oxidation of HRas cysteine thiols by metabolic stress prevents palmitoylation in vivo and contributes to endothelial cell apoptosis.
abstract: Here we demonstrate a new paradigm in redox signaling, whereby oxidants resulting from metabolic stress directly alter protein palmitoylation by oxidizing reactive cysteine thiolates. In mice fed a high-fat, high-sucrose diet and in cultured endothelial cells (ECs) treated with high palmitate and high glucose (HPHG), there was decreased HRas palmitoylation on Cys181/184 (61±24% decrease for cardiac tissue and 38±7.0% in ECs). This was due to oxidation of Cys181/184, detected using matrix-assisted laser desorption/ionization time of flight (MALDI TOF)-TOF. Decrease in HRas palmitoylation affected its compartmentalization and Ras binding domain binding activity, with a shift from plasma membrane tethering to Golgi localization. Loss of plasma membrane-bound HRas decreased growth factor-stimulated ERK phosphorylation (84±8.6% decrease) and increased apoptotic signaling (24±6.5-fold increase) after HPHG treatment that was prevented by overexpressing wild-type but not C181/184S HRas. The essential role of HRas in metabolic stress was made evident by the similar effects of expressing an inactive dominant negative N17-HRas or a MEK inhibitor. Furthermore, the relevance of thiol oxidation was demonstrated by overexpressing manganese superoxide dismutase, which improved HRas palmitoylation and ERK phosphorylation, while lessening apoptosis in HPHG treated ECs.
authors.affiliation: Vascular Biology, Whitaker Cardiovascular Institute, Boston University School of Medicine, Boston, MA 02118, USA. jrburgoy@bu.edu
authors.forename: Joseph R
authors.initials: JR
authors.lastname: Burgoyne
authors.forename: Dagmar J
authors.initials: DJ
authors.lastname: Haeussler
authors.forename: Vikas
authors.initials: V
authors.lastname: Kumar
authors.forename: Yuhan
authors.initials: Y
authors.lastname: Ji
authors.forename: David R
authors.initials: DR
authors.lastname: Pimental
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Zee
authors.forename: Cathrine E
authors.initials: CE
authors.lastname: Costello
authors.forename: Cheng
authors.initials: C
authors.lastname: Lin
authors.forename: Mark E
authors.initials: ME
authors.lastname: McComb
authors.forename: Richard A
authors.initials: RA
authors.lastname: Cohen
authors.forename: Markus M
authors.initials: MM
authors.lastname: Bachschmid
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology; D002417:Cattle; D002478:Cells, Cultured; D003545:Cysteine / Q000737:chemistry; D059305:Diet, High-Fat / Q000009:adverse effects; D004040:Dietary Carbohydrates / Q000008:administration & dosage / Q000009:adverse effects; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism*; D005947:Glucose / Q000008:administration & dosage / Q000009:adverse effects; D054878:Lipoylation; D051379:Mice; D008810:Mice, Inbred C57BL; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D016283:Proto-Oncogene Proteins p21(ras) / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D013312:Stress, Physiological; D013395:Sucrose / Q000008:administration & dosage / Q000009:adverse effects; D013482:Superoxide Dismutase / Q000378:metabolism
chemical_list: D004040:Dietary Carbohydrates; D050505:Mutant Proteins; D013395:Sucrose; D013482:Superoxide Dismutase; D020929:Mitogen-Activated Protein Kinase Kinases; D016283:Proto-Oncogene Proteins p21(ras); D005947:Glucose; D003545:Cysteine
references.citation: Biochemistry. 1996 Oct 15;35(41):13277-81
references.pmid: 8873592
references.citation: Biochem J. 2001 Apr 1;355(Pt 1):145-53
references.pmid: 11256959
references.citation: Science. 2005 Mar 18;307(5716):1746-52
references.pmid: 15705808
references.citation: Diabetes. 2005 Jun;54(6):1838-45
references.pmid: 15919807
references.citation: Antioxid Redox Signal. 2010 Oct 1;13(7):1023-32
references.pmid: 20392170
references.citation: Proc Natl Acad Sci U S A. 2003 Apr 15;100(8):4772-7
references.pmid: 12684535
references.citation: J Biol Chem. 2005 Sep 2;280(35):31141-8
references.pmid: 16000296
references.citation: Proc Natl Acad Sci U S A. 1998 Mar 3;95(5):2498-502
references.pmid: 9482914
references.citation: Cell Signal. 2003 May;15(5):463-9
references.pmid: 12639709
references.citation: J Biol Chem. 2004 Jul 9;279(28):29857-62
references.pmid: 15123696
references.citation: EMBO Rep. 2004 Nov;5(11):1053-7
references.pmid: 15520806
references.citation: J Biol Chem. 2007 Mar 30;282(13):9777-9788
references.pmid: 17255104
references.citation: Diabetes. 2000 Nov;49(11):1939-45
references.pmid: 11078463
references.citation: Nat Rev Mol Cell Biol. 2004 Nov;5(11):875-85
references.pmid: 15520807
references.citation: Proteomics Clin Appl. 2008 Jun;2(6):823-36
references.pmid: 21136882
references.citation: Diabetes. 2001 Feb;50(2):315-21
references.pmid: 11272142
references.citation: Atherosclerosis. 2003 Nov;171(1):49-55
references.pmid: 14642405
references.citation: Circulation. 2003 Mar 18;107(10):1359-65
references.pmid: 12642354
references.citation: PLoS One. 2011 Jan 06;6(1):e15866
references.pmid: 21253588
references.citation: Mol Cell Biol. 1994 Jul;14(7):4722-30
references.pmid: 8007974
references.citation: Mol Cell Biol. 2005 Aug;25(15):6722-33
references.pmid: 16024806
references.citation: J Lipid Res. 2006 May;47(5):883-91
references.pmid: 16543601
references.citation: J Cell Biol. 2010 Oct 4;191(1):23-9
references.pmid: 20876282
references.citation: Circulation. 2002 Mar 19;105(11):1291-7
references.pmid: 11901038
references.citation: Science. 1995 Nov 24;270(5240):1326-31
references.pmid: 7481820
references.citation: Cell Growth Differ. 2001 Aug;12(8):397-408
references.pmid: 11504705
references.citation: Circulation. 2002 Feb 19;105(7):788-93
references.pmid: 11854116
references.citation: J Biol Chem. 1998 Jun 19;273(25):15830-7
references.pmid: 9624183
references.citation: J Biol Chem. 2003 May 23;278(21):19292-300
references.pmid: 12642594
references.citation: Exp Cell Res. 1999 Mar 15;247(2):495-504
references.pmid: 10066377
references.citation: Mol Cell. 2011 Jan 21;41(2):173-85
references.pmid: 21255728
references.citation: J Biol Chem. 2000 Jul 21;275(29):22037-47
references.pmid: 10801823
references.citation: Cell. 1990 Oct 5;63(1):133-9
references.pmid: 2208277
references.citation: PLoS One. 2011;6(8):e23160
references.pmid: 21826235
references.citation: FASEB J. 2006 Mar;20(3):518-20
references.pmid: 16415107
references.citation: Biochem Biophys Res Commun. 2005 Oct 14;336(1):309-15
references.pmid: 16126172
references.citation: Cancer Res. 2007 Feb 1;67(3):1193-202
references.pmid: 17283155