PINK1-dependent recruitment of Parkin to mitochondria in mitophagy

Abstract

Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1) and PARK2/Parkin mutations cause autosomal recessive forms of Parkinson's disease. Upon a loss of mitochondrial membrane potential (ΔΨ m ) in human cells, cytosolic Parkin has been reported to be recruited to mitochondria, which is followed by a stimulation of mitochondrial autophagy. Here, we show that the relocation of Parkin to mitochondria induced by a collapse of ΔΨ m relies on PINK1 expression and that overexpression of WT but not of mutated PINK1 causes Parkin translocation to mitochondria, even in cells with normal ΔΨ m . We also show that once at the mitochondria, Parkin is in close proximity to PINK1, but we find no evidence that Parkin catalyzes PINK1 ubiquitination or that PINK1 phosphorylates Parkin. However, co-overexpression of Parkin and PINK1 collapses the normal tubular mitochondrial network into mitochondrial aggregates and/or large perinuclear clusters, many of which are surrounded by autophagic vacuoles. Our results suggest that Parkin, together with PINK1, modulates mitochondrial trafficking, especially to the perinuclear region, a subcellular area associated with autophagy. Thus by impairing this process, mutations in either Parkin or PINK1 may alter mitochondrial turnover which, in turn, may cause the accumulation of defective mitochondria and, ultimately, neurodegeneration in Parkinson's disease.autophagy | Parkinson's disease | phosphatase and tensin homolog-induced putative kinase 1 T he common neurodegenerative disorder Parkinson's disease (PD) occasionally can be inherited (1, 2). Parkinson disease 6/ phosphatase and tensin homolog (PTEN)-induced putative kinase-1 (PARK6/PINK1) is among the gene products associated with familial PD (2, 3). This 581-amino acid polypeptide is localized to the mitochondria and has only a single recognized functional domain, a serine/threonine kinase with a high degree of homology to that of the Ca 2+ /calmodulin kinase family. Overexpression of WT PINK1 rescues abnormal mitochondrial morphology that has been described in Drosophila carrying Pink1 mutations (4, 5), a finding that supports the notion that the mutated allele gives rise to a loss-of-function phenotype. Loss-offunction mutations in the gene encoding PARK2/Parkin (an E3 ubiquitin ligase) also can cause an autosomal recessive form of familial PD (2, 6). Parkin is thought to operate within the same molecular pathway as PINK1 to modulate mitochondrial dynamics (4, 5, 7). This possibility is intriguing, because Parkin has been reported to be essentially cytosolic (8, 9). However, we have shown that PINK1 spans the outer mitochondrial membrane, with its kinase domain facing the cytoplasm (10). These details of PINK1 topology are relevant to the reported Parkin/PINK1 genetic interaction because they place the only known functional domain of PINK1 in the same subcellular compartment as Parkin.However, the role played by Parkin, PINK1, or both in mitochondrial dynamics is still uncertain. Perhaps, the beginning of an answer to th...

Mitochondrial respiratory chain dysfunction: Implications in neurodegeneration

“…Later studies have supported Narendra's findings showing that PINK1 acts upstream from Parkin, and that both proteins are recruited to depolarized mitochondria (Kim et al, 2008;Kawajiri et al, 2010;Matsuda et al, 2010;Vives-Bauza et al, 2010). The DJ-1 protein, already known by its antioxidant, chaperone-like and transcriptional modulator functions, has been recently proposed to act in parallel with PINK1 and…”

PINK1 as a Molecular Checkpoint in the Maintenance of Mitochondrial Function and Integrity

“…The deletion of autophagyrelated gene 5 (ATG5) or treatment with autophagy inhibitors blocks this mitochondrial degradation, further confirming the Parkin-induced mitophagy (Narendra et al, 2008). Subsequent studies discovered that PINK1 is selectively translocated to mitochondria damaged by CCCP and activates their degradation via Parkin (Matsuda et al, 2010;Narendra et al, 2010a;Vives-Bauza et al, 2010). Moreover, in CCCP-treated cells, Parkin promotes the ubiquitination of impaired mitochondria and the expression of dominant-negative ubiquitin mutants prevents subsequent mitochondrial clearance, demonstrating the functional link between Parkin, ubiquitin, and mitophagy (Geisler et al, 2010;Lee et al, 2010;Matsuda et al, 2010).…”

“…Additional biochemical analyses show that PINK1 phosphorylates Parkin and promotes its mitochondrial translocalization (Kim et al, 2008a). Further studies using mitochondrial uncoupling agents such as carbonyl cyanide m-chlorophenylhydrazone (CCCP) suggest that PINK1 selectively translocates Parkin to mitochondria with low membrane potential, confirming the exis- tence of PINK1-dependent Parkin translocation (Matsuda et al, 2010;Narendra et al, 2010a;Vives-Bauza et al, 2010). Moreover, CCCP treatment also induces the accumulation of PINK1 protein in depolarized mitochondria (Matsuda et al, 2010;Narendra et al, 2010a;Vives-Bauza et al, 2010).…”

“…Further studies using mitochondrial uncoupling agents such as carbonyl cyanide m-chlorophenylhydrazone (CCCP) suggest that PINK1 selectively translocates Parkin to mitochondria with low membrane potential, confirming the exis- tence of PINK1-dependent Parkin translocation (Matsuda et al, 2010;Narendra et al, 2010a;Vives-Bauza et al, 2010). Moreover, CCCP treatment also induces the accumulation of PINK1 protein in depolarized mitochondria (Matsuda et al, 2010;Narendra et al, 2010a;Vives-Bauza et al, 2010). Subsequent studies suggested that mitochondrial depolarization inhibits PINK1 protein degradation induced by mitochondrial protease presenilin-associated rhomboid-like protein (PARL) and promotes the accumulation of full-length PINK1 in the mitochondrial outer membrane to recruit Parkin (Jin et al, 2010).…”

E2F1-dependent miR-421 regulates mitochondrial fragmentation and myocardial infarction by targeting Pink1

“…Next, we explored how Pink1 altered mitochondrial morphology in our model. Previous reports showed that Parkin requires Pink1 for mitochondrial translocation and recruitment 21,22 . Thus, we tested whether Parkin was also recruited to the mitochondria or it was a separate pathway in our signalling axis.…”