Mitochondrial matrix phosphoproteome: Effect of extra mitochondrial calcium

Rachel K. Hopper, Stefanie Carroll, Angel M. Aponte, D. Thor Johnson, Stephanie French, Rong Fong Shen, Frank A. Witzmann, Robert A. Harris, Robert S. Balaban

Research output: Contribution to journalArticle

193 Citations (Scopus)

Abstract

Post-translational modification of mitochondrial proteins by phosphorylation or dephosphorylation plays an essential role in numerous cell signaling pathways involved in regulating energy metabolism and in mitochondrion-induced apoptosis. Here we present a phosphoproteomic screen of the mitochondrial matrix proteins and begin to establish the protein phosphorylations acutely associated with calcium ions (Ca2+) signaling in porcine heart mitochondria. Forty-five phosphorylated proteins were detected by gel electrophoresis-mass spectrometry of Pro-Q Diamond staining, while many more Pro-Q Diamond-stained proteins evaded mass spectrometry detection. Time-dependent 32P incorporation in intact mitochondria confirmed the extensive matrix protein phosphoryation and revealed the dynamic nature of this process. Classes of proteins that were detected included all of the mitochondrial respiratory chain complexes, as well as enzymes involved in intermediary metabolism, such as pyruvate dehydrogenase (PDH), citrate synthase, and acyl-CoA dehydrogenases. These data demonstrate that the phosphoproteome of the mitochondrial matrix is extensive and dynamic. Ca2+ has previously been shown to activate various dehydrogenases, promote the generation of reactive oxygen species (ROS), and initiate apoptosis via cytochrome c release. To evaluate the Ca2+ signaling network, the effects of a Ca2+ challenge sufficient to release cytochrome c were evaluated on the mitochondrial phosphoproteome. Novel Ca2+-induced dephosphorylation was observed in manganese superoxide dismutase (MnSOD) as well as the previously characterized PDH. A Ca2+ dose-dependent dephosphorylation of MnSOD was associated with an ∼2-fold maximum increase in activity; neither the dephosphorylation nor activity changes were induced by ROS production in the absence of Ca2+. These data demonstrate the use of a phosphoproteome screen in determining mitochondrial signaling pathways and reveal new pathways for Ca2+ modification of mitochondrial function at the level of MnSOD.

Original languageEnglish (US)
Pages (from-to)2524-2536
Number of pages13
JournalBiochemistry
Volume45
Issue number8
DOIs
StatePublished - Feb 28 2006

Fingerprint

Calcium
Mitochondria
Superoxide Dismutase
Oxidoreductases
Diamond
Mitochondrial Proteins
Cytochromes c
Phosphorylation
Proteins
Pyruvic Acid
Acyl-CoA Dehydrogenases
Reactive Oxygen Species
Mass Spectrometry
Mass spectrometry
Apoptosis
Citrate (si)-Synthase
Heart Mitochondria
Cell signaling
Post Translational Protein Processing
Electron Transport

ASJC Scopus subject areas

  • Biochemistry

Cite this

Hopper, R. K., Carroll, S., Aponte, A. M., Johnson, D. T., French, S., Shen, R. F., ... Balaban, R. S. (2006). Mitochondrial matrix phosphoproteome: Effect of extra mitochondrial calcium. Biochemistry, 45(8), 2524-2536. https://doi.org/10.1021/bi052475e

Mitochondrial matrix phosphoproteome : Effect of extra mitochondrial calcium. / Hopper, Rachel K.; Carroll, Stefanie; Aponte, Angel M.; Johnson, D. Thor; French, Stephanie; Shen, Rong Fong; Witzmann, Frank A.; Harris, Robert A.; Balaban, Robert S.

In: Biochemistry, Vol. 45, No. 8, 28.02.2006, p. 2524-2536.

Research output: Contribution to journalArticle

Hopper, RK, Carroll, S, Aponte, AM, Johnson, DT, French, S, Shen, RF, Witzmann, FA, Harris, RA & Balaban, RS 2006, 'Mitochondrial matrix phosphoproteome: Effect of extra mitochondrial calcium', Biochemistry, vol. 45, no. 8, pp. 2524-2536. https://doi.org/10.1021/bi052475e
Hopper RK, Carroll S, Aponte AM, Johnson DT, French S, Shen RF et al. Mitochondrial matrix phosphoproteome: Effect of extra mitochondrial calcium. Biochemistry. 2006 Feb 28;45(8):2524-2536. https://doi.org/10.1021/bi052475e
Hopper, Rachel K. ; Carroll, Stefanie ; Aponte, Angel M. ; Johnson, D. Thor ; French, Stephanie ; Shen, Rong Fong ; Witzmann, Frank A. ; Harris, Robert A. ; Balaban, Robert S. / Mitochondrial matrix phosphoproteome : Effect of extra mitochondrial calcium. In: Biochemistry. 2006 ; Vol. 45, No. 8. pp. 2524-2536.
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