Insight into the Interactome of Intramitochondrial PKA Using Biotinylation-Proximity Labeling.
Biotinylation
/ methods
Cell Respiration
/ physiology
Cross-Linking Reagents
/ chemistry
Cyclic AMP-Dependent Protein Kinases
/ chemistry
HEK293 Cells
HeLa Cells
Humans
Immunoprecipitation
/ methods
Microscopy, Fluorescence
Mitochondria
/ chemistry
Mitochondrial Proteins
/ chemistry
Protein Binding
Protein Interaction Mapping
/ methods
Protein Processing, Post-Translational
Signal Transduction
Staining and Labeling
/ methods
BioID2
TIM44
mitochondria
protein kinase A
proteomics
serine/threonine phosphoprediction
Journal
International journal of molecular sciences
ISSN: 1422-0067
Titre abrégé: Int J Mol Sci
Pays: Switzerland
ID NLM: 101092791
Informations de publication
Date de publication:
05 Nov 2020
05 Nov 2020
Historique:
received:
01
10
2020
revised:
30
10
2020
accepted:
02
11
2020
entrez:
10
11
2020
pubmed:
11
11
2020
medline:
9
3
2021
Statut:
epublish
Résumé
Mitochondria are fully integrated in cell signaling. Reversible phosphorylation is involved in adjusting mitochondrial physiology to the cellular needs. Protein kinase A (PKA) phosphorylates several substrates present at the external surface of mitochondria to maintain cellular homeostasis. However, few targets of PKA located inside the organelle are known. The aim of this work was to characterize the impact and the interactome of PKA located inside mitochondria. Our results show that the overexpression of intramitochondrial PKA decreases cellular respiration and increases superoxide levels. Using proximity-dependent biotinylation, followed by LC-MS/MS analysis and in silico phospho-site prediction, we identified 21 mitochondrial proteins potentially targeted by PKA. We confirmed the interaction of PKA with TIM44 using coimmunoprecipitation and observed that TIM44-S80 is a key residue for the interaction between the protein and the kinase. These findings provide insights into the interactome of intramitochondrial PKA and suggest new potential mechanisms in the regulation of mitochondrial functions.
Identifiants
pubmed: 33167377
pii: ijms21218283
doi: 10.3390/ijms21218283
pmc: PMC7663848
pii:
doi:
Substances chimiques
Cross-Linking Reagents
0
Mitochondrial Proteins
0
Cyclic AMP-Dependent Protein Kinases
EC 2.7.11.11
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
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