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Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin
The RBR E3 ligase parkin is recruited to the outer mitochondrial membrane (OMM) during oxidative stress where it becomes activated and ubiquitinates numerous proteins. Parkin activation involves binding of a phosphorylated ubiquitin (pUb), followed by phosphorylation of the Ubl domain in parkin, bot...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Portland Press Ltd.
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9022993/ https://www.ncbi.nlm.nih.gov/pubmed/35262643 http://dx.doi.org/10.1042/BCJ20210741 |
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author | Dunkerley, Karen M. Rintala-Dempsey, Anne C. Salzano, Giulia Tadayon, Roya Hadi, Dania Barber, Kathryn R. Walden, Helen Shaw, Gary S. |
author_facet | Dunkerley, Karen M. Rintala-Dempsey, Anne C. Salzano, Giulia Tadayon, Roya Hadi, Dania Barber, Kathryn R. Walden, Helen Shaw, Gary S. |
author_sort | Dunkerley, Karen M. |
collection | PubMed |
description | The RBR E3 ligase parkin is recruited to the outer mitochondrial membrane (OMM) during oxidative stress where it becomes activated and ubiquitinates numerous proteins. Parkin activation involves binding of a phosphorylated ubiquitin (pUb), followed by phosphorylation of the Ubl domain in parkin, both mediated by the OMM kinase, PINK1. How an OMM protein is selected for ubiquitination is unclear. Parkin targeted OMM proteins have little structural or sequence similarity, with the commonality between substrates being proximity to the OMM. Here, we used chimeric proteins, tagged with ubiquitin (Ub), to evaluate parkin ubiquitination of mitochondrial acceptor proteins pre-ligated to Ub. We find that pUb tethered to the mitochondrial target proteins, Miro1 or CISD1, is necessary for parkin recruitment and essential for target protein ubiquitination. Surprisingly, phosphorylation of parkin is not necessary for the ubiquitination of either Miro1 or CISD1. Thus, parkin lacking its Ubl domain efficiently ubiquitinates a substrate tethered to pUb. Instead, phosphorylated parkin appears to stimulate free Ub chain formation. We also demonstrate that parkin ubiquitination of pUb-tethered substrates occurs on the substrate, rather than the pUb modification. We propose divergent parkin mechanisms whereby parkin-mediated ubiquitination of acceptor proteins is driven by binding to pre-existing pUb on the OMM protein and subsequent parkin phosphorylation triggers free Ub chain formation. This finding accounts for the broad spectrum of OMM proteins ubiquitinated by parkin and has implications on target design for therapeutics. |
format | Online Article Text |
id | pubmed-9022993 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Portland Press Ltd. |
record_format | MEDLINE/PubMed |
spelling | pubmed-90229932022-05-03 Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin Dunkerley, Karen M. Rintala-Dempsey, Anne C. Salzano, Giulia Tadayon, Roya Hadi, Dania Barber, Kathryn R. Walden, Helen Shaw, Gary S. Biochem J Molecular Interactions The RBR E3 ligase parkin is recruited to the outer mitochondrial membrane (OMM) during oxidative stress where it becomes activated and ubiquitinates numerous proteins. Parkin activation involves binding of a phosphorylated ubiquitin (pUb), followed by phosphorylation of the Ubl domain in parkin, both mediated by the OMM kinase, PINK1. How an OMM protein is selected for ubiquitination is unclear. Parkin targeted OMM proteins have little structural or sequence similarity, with the commonality between substrates being proximity to the OMM. Here, we used chimeric proteins, tagged with ubiquitin (Ub), to evaluate parkin ubiquitination of mitochondrial acceptor proteins pre-ligated to Ub. We find that pUb tethered to the mitochondrial target proteins, Miro1 or CISD1, is necessary for parkin recruitment and essential for target protein ubiquitination. Surprisingly, phosphorylation of parkin is not necessary for the ubiquitination of either Miro1 or CISD1. Thus, parkin lacking its Ubl domain efficiently ubiquitinates a substrate tethered to pUb. Instead, phosphorylated parkin appears to stimulate free Ub chain formation. We also demonstrate that parkin ubiquitination of pUb-tethered substrates occurs on the substrate, rather than the pUb modification. We propose divergent parkin mechanisms whereby parkin-mediated ubiquitination of acceptor proteins is driven by binding to pre-existing pUb on the OMM protein and subsequent parkin phosphorylation triggers free Ub chain formation. This finding accounts for the broad spectrum of OMM proteins ubiquitinated by parkin and has implications on target design for therapeutics. Portland Press Ltd. 2022-03-28 /pmc/articles/PMC9022993/ /pubmed/35262643 http://dx.doi.org/10.1042/BCJ20210741 Text en © 2022 The Author(s) https://creativecommons.org/licenses/by/4.0/This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY) (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Molecular Interactions Dunkerley, Karen M. Rintala-Dempsey, Anne C. Salzano, Giulia Tadayon, Roya Hadi, Dania Barber, Kathryn R. Walden, Helen Shaw, Gary S. Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
title | Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
title_full | Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
title_fullStr | Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
title_full_unstemmed | Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
title_short | Distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
title_sort | distinct phosphorylation signals drive acceptor versus free ubiquitin chain targeting by parkin |
topic | Molecular Interactions |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9022993/ https://www.ncbi.nlm.nih.gov/pubmed/35262643 http://dx.doi.org/10.1042/BCJ20210741 |
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