Mutations in the PARKIN gene are the most common cause of hereditary parkinsonism. The parkin protein comprises an N-terminal ubiquitin-like domain, a linker region containing caspase cleavage sites, a unique domain in the central portion, and a special zinc finger configuration termed RING-IBR-RING. Parkin has E3 ubiquitin-protein ligase activity and is believed to mediate proteasomal degradation of aggregation-prone proteins. Whereas the effects of mutations on the structure and function of parkin have been intensely studied, post-translational modifications of parkin and the regulation of its enzymatic activity are poorly understood. Here we report that parkin is phosphorylated both in human embryonic kidney HEK293 cells and human neuroblastoma SH-SY5Y cells. The turnover of parkin phosphorylation was rapid, because inhibition of phosphatases with okadaic acid was necessary to stabilize phosphoparkin. Phosphoamino acid analysis revealed that phosphorylation occurred mainly on serine residues under these conditions. At least five phosphorylation sites were identified, including Ser(101), Ser(131), and Ser(136) (located in the linker region) as well as Ser(296) and Ser(378) (located in the RING-IBR-RING motif). Casein kinase-1, protein kinase X and protein kinase C phosphorylated parkin in vitro, and inhibition of casein kinase-1 caused a dramatic reduction of parkin phosphorylation in cell lysates. Induction of protein folding stress in cells reduced parkin phosphorylation, and unphosphorylated parkin had slightly but significantly elevated autoubiquitination activity. Thus, complex regulation of the phosphorylation state of parkin may contribute to the unfolded protein response in stressed cells.
机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Hyun, DH
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Lee, M
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Lee, M
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Hattori, N
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Hattori, N
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Kubo, SI
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Kubo, SI
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Mizuno, Y
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Mizuno, Y
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Halliwell, B
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Halliwell, B
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Jenner, P
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Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, EnglandKings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Hyun, DH
;
Lee, M
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h-index: 0
机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Lee, M
;
Hattori, N
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Hattori, N
;
Kubo, SI
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Kubo, SI
;
Mizuno, Y
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机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Mizuno, Y
;
Halliwell, B
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h-index: 0
机构:Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England
Halliwell, B
;
Jenner, P
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h-index: 0
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Kings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, EnglandKings Coll London, Wolfson Ctr Age Related Dis, GKT, Sch Biomed Sci, London SE1 1UL, England