title: Mutant Atp13a2 proteins involved in parkinsonism are degraded by ER-associated degradation and sensitize cells to ER-stress induced cell death.
abstract: Mutations in ATP13A2 (PARK9) have been linked to juvenile parkinsonism with dementia or Kufor-Rakeb syndrome (KRS). The ATP13A2 gene encodes at least three protein isoforms that arise by alternate splicing. A previous study indicated the Atp13a2(Isoform-1) protein is localized to lysosomes, whereas three separate mutations involved in disease cause retention of the protein in the ER. One speculation is that the mutant Atp13a2(Isoform-1) proteins are misfolded and eliminated by the ER-associated degradation pathway (ERAD), which involves the dislocation of proteins from the ER to the cytoplasm for proteasome degradation. We examined whether Atp13a2 proteins are degraded by ERAD and whether the Atp13a2(Isoform-3) protein has similar localization to the Atp13a2(Isoform-1) protein. Through analysis of protein turnover and by disrupting different steps in the ERAD pathway we demonstrate that mutant Atp13a2(Isoform-1) proteins are indeed eliminated by ERAD. Thus, siRNA-mediated knockdown of erasin, a platform for assembly of an ERAD complex, or expression of a dominant negative form of p97/VCP, a protein essential for dislocation of ERAD substrates, or inhibition of the proteasome all slowed degradation of the mutant Atp13a2(Isoform-1) proteins, but not the wild-type Atp13a2(Isoform-1) protein. Immunoprecipitation assays confirmed that the Atp13a2 proteins are ubiquitinated in accord with degradation by ERAD. In contrast to Atp13a2(Isoform-1), we show Atp13a2(Isoform-3) is localized to the ER and rapidly degraded. Lastly, we show Atp13a2 mutants have increased cytotoxicity and predispose cells to ER-stress-induced cell death. These results provide new insight into the properties of wild-type and mutant Atp13a2 proteins involved in KRS.
authors.affiliation: Biochemistry and Molecular Biology Graduate Program, University of Maryland, Baltimore, 725 West Lombard Street, Baltimore, MD 21201, USA.
authors.forename: Janet
authors.initials: J
authors.lastname: Ugolino
authors.forename: Shengyun
authors.initials: S
authors.lastname: Fang
authors.forename: Christian
authors.initials: C
authors.lastname: Kubisch
authors.forename: Mervyn J
authors.initials: MJ
authors.lastname: Monteiro
mesh_terms: D016923:Cell Death; D002460:Cell Line; D059865:Endoplasmic Reticulum Stress; D060746:Endoplasmic Reticulum-Associated Degradation; D006801:Humans; D009154:Mutation; D020734:Parkinsonian Disorders / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology*; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000235:genetics* / Q000378:metabolism*
chemical_list: C514983:ATP13A2 protein, human; D046988:Proteasome Endopeptidase Complex; D006180:Proton-Translocating ATPases