RESUMO
Machado-Joseph disease (MJD), the most common dominantly inherited ataxia worldwide, is caused by a polyglutamine (polyQ) expansion in the deubiquitinating (DUB) enzyme ataxin-3. Interestingly, MJD can present clinically with features of Parkinsonism. In this study, we identify parkin, an E3 ubiquitin-ligase responsible for a common familial form of Parkinson's disease, as a novel ataxin-3 binding partner. The interaction between ataxin-3 and parkin is direct, involves multiple domains and is greatly enhanced by parkin self-ubiquitination. Moreover, ataxin-3 deubiquitinates parkin directly in vitro and in cells. Compared with wild-type ataxin-3, MJD-linked polyQ-expanded mutant ataxin-3 is more active, possibly owing to its greater efficiency at DUB K27- and K29-linked Ub conjugates on parkin. Remarkably, mutant but not wild-type ataxin-3 promotes the clearance of parkin via the autophagy pathway. The finding is consistent with the reduction in parkin levels observed in the brains of transgenic mice over-expressing polyQ-expanded but not wild-type ataxin-3, raising the intriguing possibility that increased turnover of parkin may contribute to the pathogenesis of MJD and help explain some of its parkinsonian features.
Assuntos
Doença de Machado-Joseph/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Animais , Ataxina-3 , Autofagia/genética , Células HEK293 , Humanos , Doença de Machado-Joseph/metabolismo , Camundongos , Proteínas Mutantes/genética , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/genética , Peptídeos/metabolismo , Estabilidade Proteica , Proteínas Repressoras/genética , TransfecçãoRESUMO
Mutations in the parkin gene are responsible for a common familial form of Parkinson's disease. As parkin encodes an E3 ubiquitin ligase, defects in proteasome-mediated protein degradation are believed to have a central role in the pathogenesis of Parkinson's disease. Here, we report a novel role for parkin in a proteasome-independent ubiquitination pathway. We have identified a regulated interaction between parkin and Eps15, an adaptor protein that is involved in epidermal growth factor (EGF) receptor (EGFR) endocytosis and trafficking. Treatment of cells with EGF stimulates parkin binding to both Eps15 and the EGFR and promotes parkin-mediated ubiquitination of Eps15. Binding of the parkin ubiquitin-like (Ubl) domain to the Eps15 ubiquitin-interacting motifs (UIMs) is required for parkin-mediated Eps15 ubiquitination. Furthermore, EGFR endocytosis and degradation are accelerated in parkin-deficient cells, and EGFR signalling via the phosphoinositide 3-kinase (PI(3)K)-Akt pathway is reduced in parkin knockout mouse brain. We propose that by ubiquitinating Eps15, parkin interferes with the ability of the Eps15 UIMs to bind ubiquitinated EGFR, thereby delaying EGFR internalization and degradation, and promoting PI(3)K-Akt signalling. Considering the role of Akt in neuronal survival, our results have broad new implications for understanding the pathogenesis of Parkinson's disease.
Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Receptores ErbB/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Western Blotting , Células COS , Linhagem Celular , Chlorocebus aethiops , Endocitose/efeitos dos fármacos , Endocitose/fisiologia , Fator de Crescimento Epidérmico/farmacologia , Células HeLa , Humanos , Imunoprecipitação , Camundongos , Camundongos Knockout , Células NIH 3T3 , Ligação Proteica/efeitos dos fármacos , Ligação Proteica/fisiologia , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/fisiologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Transfecção , Ubiquitina-Proteína Ligases/genéticaRESUMO
Loss-of-function mutations in the parkin gene, which encodes an E3 ubiquitin ligase, are the major cause of early-onset Parkinson's disease (PD). Decreases in parkin activity may also contribute to neurodegeneration in sporadic forms of PD. Here, we show that bcl-2-associated athanogene 5 (BAG5), a BAG family member, directly interacts with parkin and the chaperone Hsp70. Within this complex, BAG5 inhibits both parkin E3 ubiquitin ligase activity and Hsp70-mediated refolding of misfolded proteins. BAG5 enhances parkin sequestration within protein aggregates and mitigates parkin-dependent preservation of proteasome function. Finally, BAG5 enhances dopamine neuron death in an in vivo model of PD, whereas a mutant that inhibits BAG5 activity attenuates dopaminergic neurodegeneration. This contrasts with the antideath functions ascribed to BAG family members and suggests a potential role for BAG5 in promoting neurodegeneration in sporadic PD through its functional interactions with parkin and Hsp70.