PMID- 28442264 OWN - NLM STAT- MEDLINE DCOM- 20171027 LR - 20231112 IS - 0005-2728 (Print) IS - 1878-2434 (Electronic) IS - 0005-2728 (Linking) VI - 1858 IP - 7 DP - 2017 Jul TI - Mitochondrial LON protease-dependent degradation of cytochrome c oxidase subunits under hypoxia and myocardial ischemia. PG - 519-528 LID - S0005-2728(17)30059-2 [pii] LID - 10.1016/j.bbabio.2017.04.003 [doi] AB - The mitochondrial ATP dependent matrix protease, Lon, is involved in the maintenance of mitochondrial DNA nucleoids and degradation of abnormal or misfolded proteins. The Lon protease regulates mitochondrial Tfam (mitochondrial transcription factor A) level and thus modulates mitochondrial DNA (mtDNA) content. We have previously shown that hypoxic stress induces the PKA-dependent phosphorylation of cytochrome c oxidase (CcO) subunits I, IVi1, and Vb and a time-dependent reduction of these subunits in RAW 264.7 murine macrophages subjected to hypoxia and rabbit hearts subjected to ischemia/reperfusion. Here, we show that Lon is involved in the preferential turnover of phosphorylated CcO subunits under hypoxic/ischemic stress. Induction of Lon protease occurs at 6 to 12 h of hypoxia and this increase coincides with lower CcO subunit contents. Over-expression of flag-tagged wild type and phosphorylation site mutant Vb and IVi1 subunits (S40A and T52A, respectively) caused marked degradation of wild type protein under hypoxia while the mutant proteins were relatively resistant. Furthermore, the recombinant purified Lon protease degraded the phosphorylated IVi1 and Vb subunits, while the phosphorylation-site mutant proteins were resistant to degradation. 3D structural modeling shows that the phosphorylation sites are exposed to the matrix compartment, accessible to matrix PKA and Lon protease. Hypoxic stress did not alter CcO subunit levels in Lon depleted cells, confirming its role in CcO turnover. Our results therefore suggest that Lon preferentially degrades the phosphorylated subunits of CcO and plays a role in the regulation of CcO activity in hypoxia and ischemia/reperfusion injury. CI - Copyright (c) 2017 Elsevier B.V. All rights reserved. FAU - Sepuri, Naresh B V AU - Sepuri NBV AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. FAU - Angireddy, Rajesh AU - Angireddy R AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. FAU - Srinivasan, Satish AU - Srinivasan S AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. FAU - Guha, Manti AU - Guha M AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. FAU - Spear, Joseph AU - Spear J AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. FAU - Lu, Bin AU - Lu B AD - Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers The State University, New Jersey Medical School, 225 Warren Street, Newark, NJ 17103-3535, USA. FAU - Anandatheerthavarada, Hindupur K AU - Anandatheerthavarada HK AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. FAU - Suzuki, Carolyn K AU - Suzuki CK AD - Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers The State University, New Jersey Medical School, 225 Warren Street, Newark, NJ 17103-3535, USA. FAU - Avadhani, Narayan G AU - Avadhani NG AD - Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce Street, Philadelphia, PA 19104-6009, USA. Electronic address: narayan@vet.upenn.edu. LA - eng GR - R01 AA022986/AA/NIAAA NIH HHS/United States GR - R01 AR067066/AR/NIAMS NIH HHS/United States GR - R01 GM034883/GM/NIGMS NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, Non-U.S. Gov't DEP - 20170423 PL - Netherlands TA - Biochim Biophys Acta Bioenerg JT - Biochimica et biophysica acta. Bioenergetics JID - 101731706 RN - 0 (Mitochondrial Proteins) RN - 0 (Protein Subunits) RN - 0 (RNA, Small Interfering) RN - 0 (Recombinant Proteins) RN - EC 1.9.3.1 (Electron Transport Complex IV) RN - EC 2.7.11.11 (Cyclic AMP-Dependent Protein Kinases) RN - EC 3.4.21.- (ATP-Dependent Proteases) RN - EC 3.4.21.- (LONP1 protein, human) RN - EC 3.4.21.53 (LONP1 protein, mouse) SB - IM MH - ATP-Dependent Proteases/chemistry/genetics/*metabolism MH - Animals MH - Cell Hypoxia/*physiology MH - Cyclic AMP-Dependent Protein Kinases/metabolism MH - Electron Transport Complex IV/*metabolism MH - Humans MH - Male MH - Mice MH - Mitochondria, Heart/*enzymology MH - Mitochondrial Proteins/chemistry/genetics/*metabolism MH - Models, Molecular MH - Myocardial Ischemia/*enzymology MH - Phosphorylation MH - Protein Conformation MH - Protein Processing, Post-Translational MH - Protein Subunits MH - RAW 264.7 Cells MH - RNA Interference MH - RNA, Small Interfering/genetics MH - Rabbits MH - Recombinant Proteins/metabolism PMC - PMC5507603 MID - NIHMS876395 OTO - NOTNLM OT - 3D modeling OT - CcO subunits OT - Heart ischemia OT - Hypoxia OT - Mitochondrial LON OT - PKA dependent phosphorylation COIS- Conflicts of interest: Authors declare no conflict of interest with any of the funding agencies. EDAT- 2017/04/27 06:00 MHDA- 2017/10/28 06:00 PMCR- 2018/07/01 CRDT- 2017/04/27 06:00 PHST- 2016/06/01 00:00 [received] PHST- 2017/04/17 00:00 [revised] PHST- 2017/04/21 00:00 [accepted] PHST- 2017/04/27 06:00 [pubmed] PHST- 2017/10/28 06:00 [medline] PHST- 2017/04/27 06:00 [entrez] PHST- 2018/07/01 00:00 [pmc-release] AID - S0005-2728(17)30059-2 [pii] AID - 10.1016/j.bbabio.2017.04.003 [doi] PST - ppublish SO - Biochim Biophys Acta Bioenerg. 2017 Jul;1858(7):519-528. doi: 10.1016/j.bbabio.2017.04.003. Epub 2017 Apr 23.