Description
The inner mitochondrial membrane protein MPV17 has been described to be involved in rare human mitochondrial diseases associated with mitochondrial DNA (mtDNA) maintenance defects. However, the mechanistic link between MPV17 and mtDNA maintenance is still lacking even though several functions have been suggested for this protein. Indeed, MPV17 has been proposed to act as a non-selective channel aiming at maintaining the inner mitochondrial membrane potential under stress. It has also been proposed to be involved in the nucleotide salvage pathway thanks to deoxynucleotide import into the mitochondrial matrix. However, the function of MPV17 seems to be related to its association with a >600 kDa complex, as functionally described in yeast and mouse models. Therefore, in order to get new clues about MPV17 function, we decided to characterize the MPV17 proxisome using BioID. This proximity labeling assay is based on the biotinylating activity of an engineered biotin ligase BirA*. Upon fusion with a protein of interest, it allows for the biotinylation of the surrounding proteins in a 10 nm range. Those proteins can then be purified using streptavidin beads and identified using mass spectrometry.MPV17 was fused with the BirA* enzyme and, using a matrixial biotin ligase as control, a label-free enrichment analysis by mass spectrometry was performed. The Gene Ontology analysis of the BioID results indicated a highly significant association of MPV17 with the MItochondrial contact site and Cristae Organizing System (MICOS) complex and more generally with the Mitochondrial Intermembrane space Bridging (MIB) supercomplex. The MICOS complex is involved in the cristae formation and cristae junction stabilization. Thanks to its interaction with the outer membrane complex SAM50, it bridges the outer and inner membranes of mitochondria and ensures mitochondria ultrastructure maintenance. The disruption of core components of the MICOS complex leads to cristae disruption and ultimately to mtDNA nucleoid destabilization, impaired mtDNA replication and mtDNA depletion. In addition, this association of MPV17 with the MICOS complex is in line with the mitochondrial ultrastructure defects observed in MPV17-deficient yeast and mouse models. However, even though we confirmed by immunoprecipitation experiments that MPV17 does interact with the MICOS, our preliminary mitochondrial ultrastructure analyses of MPV17-KO HEK293T do not support a role for MPV17 in cristae formation or maintenance.
However, other candidates were also found close to MPV17 including the Exonuclease G, involved in mitochondrial long-patch base excision repair, the catalytic subunit of the DNA-PK, involved in the nuclear DNA Damage Response, and ATAD3B, a mitophagy receptor. Both Exonuclease G and DNA-PKcs are associated with DNA damage sensing and repair whereas ATAD3B is involved in the clearance of mtDNA damages and is one of the most significantly enriched candidates in the MPV17 proxisome. Therefore, our current working hypothesis is that MPV17 might be associated with mtDNA quality control and mtDNA damage sensing. This hypothesis is supported by the stronger mitophagy activity observed in MPV17-KO cells. Ultimately, this study will bring new clues about MPV17 function, possibly paving the way for the identification of new targets for a potential treatment that is still lacking up to now.
| Période | 15 mai 2022 → 19 mai 2022 |
|---|---|
| Titre de l'événement | EMBO Workshop on Molecular Biology of Mitochondrial Gene Maintenance and Expression: Potentiation of radiotherapy through mitophagy inhibition and induced oxidative stress |
| Type d'événement | Colloque |
| Lieu | Bro, SuèdeAfficher sur la carte |
| Niveau de reconnaissance | International |