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Differential submitochondrial localization of PINK1 as a molecular switch for mediating distinct mitochondrial signaling pathways.

ABSTRACT: Mutations in mitochondrial kinase PINK1 cause Parkinson disease (PD), but the submitochondrial site(s) of PINK1 action remains unclear. Here, we report that three-dimensional structured illumination microscopy (3D-SIM) enables super-resolution imaging of protein submitochondrial localization. Dual-color 3D-SIM imaging analysis revealed that PINK1 resides in the cristae membrane and intracristae space but not on the outer mitochondrial membrane (OMM) of healthy mitochondria. Under normal physiological conditions, PINK1 colocalizes with its substrate TRAP1 in the cristae membrane and intracristae space. In response to mitochondrial depolarization, PINK1, but not TRAP1, translocates to the OMM. The PINK1 translocation to the OMM of depolarized mitochondria is independent of new protein synthesis and requires combined action of PINK1 transmembrane domain and C-terminal region. We found that mitochondrial depolarization-induced PINK1 OMM translocation is required for recruitment of parkin to the OMM of damaged mitochondria. Our findings suggest that differential submitochondrial localization of PINK1 serves as a molecular switch for mediating two distinct mitochondrial signaling pathways in maintenance of mitochondrial homeostasis. Furthermore, our study provides evidence for the involvement of deregulated PINK1 submitochondrial localization in PD pathogenesis.

SUBMITTER: Fallaize D

PROVIDER: S-EPMC4684445 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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Differential submitochondrial localization of PINK1 as a molecular switch for mediating distinct mitochondrial signaling pathways.

Fallaize Dana D Chin Lih-Shen LS Li Lian L

Cellular signalling 20151006 12

Mutations in mitochondrial kinase PINK1 cause Parkinson disease (PD), but the submitochondrial site(s) of PINK1 action remains unclear. Here, we report that three-dimensional structured illumination microscopy (3D-SIM) enables super-resolution imaging of protein submitochondrial localization. Dual-color 3D-SIM imaging analysis revealed that PINK1 resides in the cristae membrane and intracristae space but not on the outer mitochondrial membrane (OMM) of healthy mitochondria. Under normal physiolo ...[more]

PMID: 26436374

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Project description:Mitochondria are highly dynamic organelles with strict quality control processes that maintain cellular homeostasis. Within axons, coordinated cycles of fission-fusion mediated by dynamin related GTPase protein (DRP1) and mitofusins (MFN), together with regulated motility of healthy mitochondria anterogradely and damaged/oxidized mitochondria retrogradely, control mitochondrial shape, distribution and size. Disruption of this tight regulation has been linked to aberrant oxidative stress and mitochondrial dysfunction causing mitochondrial disease and neurodegeneration. Although pharmacological induction of Parkinson's disease (PD) in humans/animals with toxins or in mice overexpressing α-synuclein (α-syn) exhibited mitochondrial dysfunction and oxidative stress, mice lacking α-syn showed resistance to mitochondrial toxins; yet, how α-syn influences mitochondrial dynamics and turnover is unclear. Here, we isolate the mechanistic role of α-syn in mitochondrial homeostasis in vivo in a humanized Drosophila model of Parkinson's disease (PD). We show that excess α-syn causes fragmented mitochondria, which persists with either truncation of the C-terminus (α-syn1-120) or deletion of the NAC region (α-synΔNAC). Using in vivo oxidation reporters Mito-roGFP2-ORP1/GRX1 and MitoTimer, we found that α-syn-mediated fragments were oxidized/damaged, but α-syn1-120-induced fragments were healthy, suggesting that the C-terminus is required for oxidation. α-syn-mediated oxidized fragments showed biased retrograde motility, but α-syn1-120-mediated healthy fragments did not, demonstrating that the C-terminus likely mediates the retrograde motility of oxidized mitochondria. Depletion/inhibition or excess DRP1-rescued α-syn-mediated fragmentation, oxidation, and the biased retrograde motility, indicating that DRP1-mediated fragmentation is likely upstream of oxidation and motility changes. Further, excess PINK/Parkin, two PD-associated proteins that function to coordinate mitochondrial turnover via induction of selective mitophagy, rescued α-syn-mediated membrane depolarization, oxidation and cell death in a C-terminus-dependent manner, suggesting a functional interaction between α-syn and PINK/Parkin. Taken together, our findings identify distinct roles for α-syn in mitochondrial homeostasis, highlighting a previously unknown pathogenic pathway for the initiation of PD.

Project description:The proteoglycan agrin is required for postsynaptic differentiation at the skeletal neuromuscular junction, but is also associated with basal laminae in numerous other tissues, and with the surfaces of some neurons. Little is known about its roles at sites other than the neuromuscular junction, or about how its expression and subcellular localization are regulated in any tissue. Here we demonstrate that the murine agrin gene generates two proteins with different NH(2) termini, and present evidence that these isoforms differ in subcellular localization, tissue distribution, and function. The two isoforms share approximately 1,900 amino acids (aa) of common sequence following unique NH(2) termini of 49 or 150 aa; we therefore call them short NH(2)-terminal (SN) and long NH(2)-terminal (LN) isoforms. In the mouse genome, LN-specific exons are upstream of an SN-specific exon, which is in turn upstream of common exons. LN-agrin is expressed in both neural and nonneural tissues. In spinal cord it is expressed in discrete subsets of cells, including motoneurons. In contrast, SN-agrin is selectively expressed in the nervous system but is widely distributed in many neuronal cell types. Both isoforms are externalized from cells but LN-agrin assembles into basal laminae whereas SN-agrin remains cell associated. Differential expression of the two isoforms appears to be transcriptionally regulated, whereas the unique SN and LN sequences direct their distinct subcellular localizations. Insertion of a "gene trap" construct into the mouse genome between the LN and SN exons abolished expression of LN-agrin with no detectable effect on expression levels of SN-agrin or on SN-agrin bioactivity in vitro. Agrin protein was absent from all basal laminae in mice lacking LN-agrin transcripts. The formation of the neuromuscular junctions was as drastically impaired in these mutants as in mice lacking all forms of agrin. Thus, basal lamina-associated LN-agrin is required for neuromuscular synaptogenesis, whereas cell-associated SN-agrin may play distinct roles in the central nervous system.

Dataset Information

Differential submitochondrial localization of PINK1 as a molecular switch for mediating distinct mitochondrial signaling pathways.

Publications

Differential submitochondrial localization of PINK1 as a molecular switch for mediating distinct mitochondrial signaling pathways.

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