Novel Insights into the PKCβ-dependent Regulation of the Oxidoreductase p66Shc.
Dysfunctional mitochondria contribute to the development of many diseases and pathological conditions through the excessive production of reactive oxygen species (ROS), and, where studied, ablation of p66Shc (p66) was beneficial. p66 translocates to the mitochondria and oxidizes cytochrome c to yield H2O2, which in turn initiates cell death. PKCβ-mediated phosphorylation of serine 36 in p66 has been implicated as a key regulatory step preceding mitochondrial translocation, ROS production, and cell death, and PKCβ thus may provide a target for therapeutic intervention. We performed a reassessment of PKCβ regulation of the oxidoreductase activity of p66. Although our experiments did not substantiate Ser36phosphorylation by PKCβ, they instead provided evidence for Ser139and Ser213as PKCβ phosphorylation sites regulating the pro-oxidant and pro-apoptotic function of p66. Mutation of another predicted PKCβ phosphorylation site also located in the phosphotyrosine binding domain, threonine 206, had no phenotype. Intriguingly, p66 with Thr206and Ser213mutated to glutamic acid showed a gain-of-function phenotype with significantly increased ROS production and cell death induction. Taken together, these data argue for a complex mechanism of PKCβ-dependent regulation of p66 activation involving Ser139and a motif surrounding Ser213.
From the Daniel Swarovski Research Laboratory, Department of Visceral, Transplant, and Thoracic Surgery.
Martina
M
Haller
From the Daniel Swarovski Research Laboratory, Department of Visceral, Transplant, and Thoracic Surgery.
Sana
S
Khalid
Division of Clinical Biochemistry, Protein Micro-Analysis Facility.
Leopold
L
Kremser
Department for Pharmacology and Genetics, Division of Translational Cell Genetics, and.
Friedrich
F
Fresser
From the Daniel Swarovski Research Laboratory, Department of Visceral, Transplant, and Thoracic Surgery.
Tobias
T
Furlan
Department for Anesthesiology and Intensive Care, Medical University of Innsbruck, 6020 Innsbruck, Austria.
Martin
M
Hermann
From the Daniel Swarovski Research Laboratory, Department of Visceral, Transplant, and Thoracic Surgery.
Julia
J
Guenther
From the Daniel Swarovski Research Laboratory, Department of Visceral, Transplant, and Thoracic Surgery.
Astrid
A
Drasche
the Biotechnology Center of Oslo, 0349 Oslo, Norway, and.
Michael
M
Leitges
the European Institute of Oncology, 20139 Milano, Italy.
Marco
M
Giorgio
Department for Pharmacology and Genetics, Division of Translational Cell Genetics, and.
Gottfried
G
Baier
Division of Clinical Biochemistry, Protein Micro-Analysis Facility.
Herbert
H
Lindner
From the Daniel Swarovski Research Laboratory, Department of Visceral, Transplant, and Thoracic Surgery, jakob.troppmair@i-med.ac.at.
Jakob
J
Troppmair
D016475:3T3 Cells; D000818:Animals; D016923:Cell Death; D017353:Gene Deletion; D057809:HEK293 Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D051379:Mice; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D018384:Oxidative Stress; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017354:Point Mutation; D064546:Protein Kinase C beta / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012694:Serine / Q000235:genetics / Q000378:metabolism; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1 / Q000235:genetics / Q000378:metabolism*
D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; C528024:SHC1 protein, human; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D012694:Serine; D006861:Hydrogen Peroxide; D010088:Oxidoreductases; D064546:Protein Kinase C beta
PKC; cell death; p66shc; phosphorylation; phosphotyrosine binding (PTB) domain; reactive oxygen species (ROS); redox signaling