Cargando…

A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development

Ubiquitination has essential roles in neuronal development and function. Ubiquitin proteomics studies on yeast and HeLa cells have proven very informative, but there still is a gap regarding neuronal tissue-specific ubiquitination. In an organism context, direct evidence for the ubiquitination of ne...

Descripción completa

Detalles Bibliográficos
Autores principales: Franco, Maribel, Seyfried, Nicholas T., Brand, Andrea H., Peng, Junmin, Mayor, Ugo
Formato: Texto
Lenguaje:English
Publicado: The American Society for Biochemistry and Molecular Biology 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3098581/
https://www.ncbi.nlm.nih.gov/pubmed/20861518
http://dx.doi.org/10.1074/mcp.M110.002188
_version_ 1782203981304430592
author Franco, Maribel
Seyfried, Nicholas T.
Brand, Andrea H.
Peng, Junmin
Mayor, Ugo
author_facet Franco, Maribel
Seyfried, Nicholas T.
Brand, Andrea H.
Peng, Junmin
Mayor, Ugo
author_sort Franco, Maribel
collection PubMed
description Ubiquitination has essential roles in neuronal development and function. Ubiquitin proteomics studies on yeast and HeLa cells have proven very informative, but there still is a gap regarding neuronal tissue-specific ubiquitination. In an organism context, direct evidence for the ubiquitination of neuronal proteins is even scarcer. Here, we report a novel proteomics strategy based on the in vivo biotinylation of ubiquitin to isolate ubiquitin conjugates from the neurons of Drosophila melanogaster embryos. We confidently identified 48 neuronal ubiquitin substrates, none of which was yet known to be ubiquitinated. Earlier proteomics and biochemical studies in non-neuronal cell types had identified orthologs to some of those but not to others. The identification here of novel ubiquitin substrates, those with no known ubiquitinated ortholog, suggests that proteomics studies must be performed on neuronal cells to identify ubiquitination pathways not shared by other cell types. Importantly, several of those newly found neuronal ubiquitin substrates are key players in synaptogenesis. Mass spectrometry results were validated by Western blotting to confirm that those proteins are indeed ubiquitinated in the Drosophila embryonic nervous system and to elucidate whether they are mono- or polyubiquitinated. In addition to the ubiquitin substrates, we also identified the ubiquitin carriers that are active during synaptogenesis. Identifying endogenously ubiquitinated proteins in specific cell types, at specific developmental stages, and within the context of a living organism will allow understanding how the tissue-specific function of those proteins is regulated by the ubiquitin system.
format Text
id pubmed-3098581
institution National Center for Biotechnology Information
language English
publishDate 2011
publisher The American Society for Biochemistry and Molecular Biology
record_format MEDLINE/PubMed
spelling pubmed-30985812011-05-27 A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development Franco, Maribel Seyfried, Nicholas T. Brand, Andrea H. Peng, Junmin Mayor, Ugo Mol Cell Proteomics Special Issue Ubiquitination has essential roles in neuronal development and function. Ubiquitin proteomics studies on yeast and HeLa cells have proven very informative, but there still is a gap regarding neuronal tissue-specific ubiquitination. In an organism context, direct evidence for the ubiquitination of neuronal proteins is even scarcer. Here, we report a novel proteomics strategy based on the in vivo biotinylation of ubiquitin to isolate ubiquitin conjugates from the neurons of Drosophila melanogaster embryos. We confidently identified 48 neuronal ubiquitin substrates, none of which was yet known to be ubiquitinated. Earlier proteomics and biochemical studies in non-neuronal cell types had identified orthologs to some of those but not to others. The identification here of novel ubiquitin substrates, those with no known ubiquitinated ortholog, suggests that proteomics studies must be performed on neuronal cells to identify ubiquitination pathways not shared by other cell types. Importantly, several of those newly found neuronal ubiquitin substrates are key players in synaptogenesis. Mass spectrometry results were validated by Western blotting to confirm that those proteins are indeed ubiquitinated in the Drosophila embryonic nervous system and to elucidate whether they are mono- or polyubiquitinated. In addition to the ubiquitin substrates, we also identified the ubiquitin carriers that are active during synaptogenesis. Identifying endogenously ubiquitinated proteins in specific cell types, at specific developmental stages, and within the context of a living organism will allow understanding how the tissue-specific function of those proteins is regulated by the ubiquitin system. The American Society for Biochemistry and Molecular Biology 2011-05 2010-09-22 /pmc/articles/PMC3098581/ /pubmed/20861518 http://dx.doi.org/10.1074/mcp.M110.002188 Text en © 2011 by The American Society for Biochemistry and Molecular Biology, Inc. Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) applies to Author Choice Articles
spellingShingle Special Issue
Franco, Maribel
Seyfried, Nicholas T.
Brand, Andrea H.
Peng, Junmin
Mayor, Ugo
A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development
title A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development
title_full A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development
title_fullStr A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development
title_full_unstemmed A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development
title_short A Novel Strategy to Isolate Ubiquitin Conjugates Reveals Wide Role for Ubiquitination during Neural Development
title_sort novel strategy to isolate ubiquitin conjugates reveals wide role for ubiquitination during neural development
topic Special Issue
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3098581/
https://www.ncbi.nlm.nih.gov/pubmed/20861518
http://dx.doi.org/10.1074/mcp.M110.002188
work_keys_str_mv AT francomaribel anovelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT seyfriednicholast anovelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT brandandreah anovelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT pengjunmin anovelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT mayorugo anovelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT francomaribel novelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT seyfriednicholast novelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT brandandreah novelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT pengjunmin novelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment
AT mayorugo novelstrategytoisolateubiquitinconjugatesrevealswideroleforubiquitinationduringneuraldevelopment