Phosphatase and Tensin Homolog (PTEN)-induced Putative Kinase 1 (PINK1)-dependent Ubiquitination of Endogenous Parkin Attenuates Mitophagy

Abstract

Background: The Parkinson disease-related proteins PINK1 and Parkin initiate mitophagy of damaged mitochondria. Results: Endogenous Parkin is not sufficient to induce mitophagy due to PINK1-dependent ubiquitination of Parkin. Conclusion: Mitophagy is detectable only with supraphysiological levels of Parkin and differs between fibroblasts and iPS-derived neurons. Significance: Stresses the importance of future studies in Parkinson disease-relevant tissue, i.e., dopaminergic neurons. Mutations in the E3 ubiquitin ligase Parkin and the mitochon-drial PTEN-induced putative kinase 1 (PINK1) have been iden-tified to cause autosomal recessive forms of familial Parkinson disease, with PINK1 functioning upstream of Parkin in a path-way important for the maintenance of mitochondrial function and morphology. Upon the loss of the mitochondrial membrane potential, Parkin translocates to mitochondria in a PINK1-de-pendent manner to ubiquitinate mitochondrial proteins. Par-kin-mediated polyubiquitination of outer mitochondrial mem-brane (OMM) proteins recruits the ubiquitin-and LC3-binding adaptor protein p62 to mitochondria and induces mitophagy. Although previous studies examined mitophagy in established cell lines through overexpression approaches, there is an impera-tive to study the role of endogenous Parkin and PINK1 in human-derived and biologically relevant cell models. Here, we demon-strate in human primary fibroblasts and induced pluripotent stem-derived neurons from controls and PINK1 mutation carriers that endogenous levels of Parkin are not sufficient to initiate mitophagy upon loss of the mitochondrial membrane potential, caused by its (self-)ubiquitination, followed by degradation via the ubiquitin proteasome system. Next, we showed differential PINK1-dependent, Parkin-mediated ubiquitination of OMM pro-teins, which is Parkin dose-dependent and affects primarily OMM proteins of higher molecular mass. In contrast to the situation fibroblasts, we did not detect mitophagy in induced pluripotent stem-derived neurons even upon overexpression of Parkin. Taken together, our data demonstrate that mitophagy differs between human non-neuronal and neuronal cells and between " endoge-nous " and " Parkin-overexpressing " cellular models.