16 Aug 2013. Growing evidence points to mitochondrial quality control as a key feature of familial Parkinson’s disease (PD) and related disorders. An August 11 Nature Neuroscience paper bolsters that notion. Researchers led by Helene Plun-Favreau at University College London, UK, connect the parkinsonism-linked gene F-box only protein 7 (Fbxo7) to mitophagy—a system cells use to dispose of damaged mitochondria. The mitochondrial kinase Pink1 and parkin, a ubiquitin ligase, drive mitophagy, and genetic mutations in each cause early-onset PD. The new research shows that Fbxo7, another ubiquitin ligase, binds parkin and helps direct it to mitochondria. This triggers mitophagy to remove dysfunctional mitochondria in neurons, Plun-Favreau said. Co-corresponding authors Heike Laman at the University of Cambridge, and Alexander Whitworth at the University of Sheffield collaborated on the study. While the findings strengthen the case for mitophagy regulators as potential therapeutic targets for familial PD, it remains unclear to what extent this process relates to the more common sporadic forms of PD.
In 2008, scientists found mutations in Fbxo7, a.k.a. PARK 15, in people with a rare form of recessive parkinsonism called parkinsonian-pyramidal disease (see Shojaee et al., 2008; Di Fonzo et al., 2009; Paisán-Ruiz et al., 2010). Their symptoms resemble those of PD patients with parkin and Pink1 mutations, but strike earlier and include spasticity due to white matter lesions (see Deng et al., 2013).
Noticing a putative mitochondrial targeting sequence at Fbxo7’s N-terminal end, co-lead author Victoria Burchell and colleagues, including co-first authors David Nelson and Alvaro Sanchez-Martinez, began exploring a mitochondrial role for the protein by testing if it interacts with parkin (a.k.a. PARK 2). The researchers overexpressed tagged versions of parkin and Fbxo7 in several human cell lines and detected each in immunoprecipitates of the other, concluding that the two proteins interact. They found that Fbxo7’s N-terminal ubiquitin-like region is necessary and sufficient for parkin binding. Both proteins reside primarily in the cytosol but appeared to associate with each other more in mitochondria, as judged by Western blotting of immunoprecipitates from cellular subfractions.
Fbxo7 also binds Pink1 (PARK 6), the scientists found. However, that association required a small stretch of Fbxo7 just downstream of the N-terminal ubiquitin-like domain needed for parkin binding, “raising the possibility that the interactions of Fbxo7, parkin, and Pink1 may be mutually exclusive or form ternary complexes,” the authors wrote.
How might mutations in these three proteins lead to PD? It is well established that parkin and Pink1 drive mitophagy, with the latter functioning upstream. Pink1 accumulates quickly on mitochondria damaged by CCCP, a compound that disrupts the organelle’s electrochemical gradient (see Narendra et al., 2010). The damage somehow nudges Pink1 onto the impaired mitochondria where it recruits parkin (see Narendra et al., 2008; Vives-Bauza et al., 2010). The current study proposes that Fbxo7 facilitates this translocation.
Compared to normal cells, Fbxo7 glommed onto parkin in mitochondria of SH-SY5Y human neuroblastoma cells that were treated with CCCP, suggesting Fbxo7 and parkin interact more readily in mitochondria on the wane. Conversely, silencing Fbxo7 kept parkin out of the damaged mitochondria. Fbxo7 knockdown also reduced ubiquitination of mitofusin 1, a GTPase on the outer mitochondrial membrane that scientists identified as a parkin substrate earlier this year (see ARF news story on Kim et al., 2013). Fbxo7 overexpression restored mitofusin ubiquitination. Patient fibroblasts homozygous for a pathogenic Fbxo7 mutation had similar abnormalities as Fbxo7-deficient neuroblastoma cells.
Work on Drosophila indicates these interactions are functionally important. Overexpression of Fbxo7 corrected motor deficiencies in Parkin-deficient flies, whereas Fbxo7 with pathogenic mutations (T22M, R378G or R498X), or lacking the mitochondrial targeting sequence, failed. It is curious that Fbxo7 rescued flies with no parkin, noted Mark Cookson of the National Institute on Aging in Bethesda, Maryland. This suggests the rescue could not result from physical interaction between Fbxo7 and parkin, wrote Cookson (see full comment below). The authors propose Fbxo7 may interact with another ubiquitin ligase.
The present findings add Fbxo7 to a growing list of parkinsonism-associated genes involved in mitochondrial dynamics and confirms the importance of mitophagy in early-onset PD-like disorders. However, Taylor noted, “the big question is whether patients with these rare familial syndromes reflect what’s happening in the more common sporadic forms of disease.” Juvenile forms make up less than a tenth of all PD and parkinsonism cases, and parkin mutations account for about 40 percent of the early-onset patients. Mutations in Pink1 cause 10 percent of early-onset cases, and Fbxo7 mutations are rarer still.
Plun-Favreau said her team plans to analyze Fbxo7 and the other mitophagy players in neurons differentiated from fibroblasts of patients with PD/parkinsonism mutations. She also hopes to collaborate with industry to find compounds that enhance parkin relocation to unhealthy mitochondria or otherwise drive mitophagy.—Esther Landhuis.
Burchell VS, Nelson DE, Sanchez-Martinez A, Delgado-Camprubi M, Ivatt RM, Pogson JH, Randle SJ, Wray S, Lewis PA, Houlden H, Abramov AY, Hardy J, Wood NW, Whitworth AJ, Laman H, Plun-Favreau H. The Parkinson’s disease-linked proteins Fbxo7 and Parkin interact to mediate mitophagy. 11 Aug 2013. Abstract