Cargando…

A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans

Endoplasmic-reticulum associated degradation (ERAD) is a major cellular misfolded protein disposal pathway that is well conserved from yeast to mammals. In yeast, a mutant of carboxypeptidase Y (CPY*) was found to be a luminal ER substrate and has served as a useful marker to help identify modifiers...

Descripción completa

Detalles Bibliográficos
Autores principales: Miedel, Mark T., Graf, Nathan J., Stephen, Kate E., Long, Olivia S., Pak, Stephen C., Perlmutter, David H., Silverman, Gary A., Luke, Cliff J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3388072/
https://www.ncbi.nlm.nih.gov/pubmed/22768338
http://dx.doi.org/10.1371/journal.pone.0040145
_version_ 1782237140789231616
author Miedel, Mark T.
Graf, Nathan J.
Stephen, Kate E.
Long, Olivia S.
Pak, Stephen C.
Perlmutter, David H.
Silverman, Gary A.
Luke, Cliff J.
author_facet Miedel, Mark T.
Graf, Nathan J.
Stephen, Kate E.
Long, Olivia S.
Pak, Stephen C.
Perlmutter, David H.
Silverman, Gary A.
Luke, Cliff J.
author_sort Miedel, Mark T.
collection PubMed
description Endoplasmic-reticulum associated degradation (ERAD) is a major cellular misfolded protein disposal pathway that is well conserved from yeast to mammals. In yeast, a mutant of carboxypeptidase Y (CPY*) was found to be a luminal ER substrate and has served as a useful marker to help identify modifiers of the ERAD pathway. Due to its ease of genetic manipulation and the ability to conduct a genome wide screen for modifiers of molecular pathways, C. elegans has become one of the preferred metazoans for studying cell biological processes, such as ERAD. However, a marker of ERAD activity comparable to CPY* has not been developed for this model system. We describe a mutant of pro-cathepsin L fused to YFP that no longer targets to the lysosome, but is efficiently eliminated by the ERAD pathway. Using this mutant pro-cathepsin L, we found that components of the mammalian ERAD system that participate in the degradation of ER luminal substrates were conserved in C. elegans. This transgenic line will facilitate high-throughput genetic or pharmacological screens for ERAD modifiers using widefield epifluorescence microscopy.
format Online
Article
Text
id pubmed-3388072
institution National Center for Biotechnology Information
language English
publishDate 2012
publisher Public Library of Science
record_format MEDLINE/PubMed
spelling pubmed-33880722012-07-05 A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans Miedel, Mark T. Graf, Nathan J. Stephen, Kate E. Long, Olivia S. Pak, Stephen C. Perlmutter, David H. Silverman, Gary A. Luke, Cliff J. PLoS One Research Article Endoplasmic-reticulum associated degradation (ERAD) is a major cellular misfolded protein disposal pathway that is well conserved from yeast to mammals. In yeast, a mutant of carboxypeptidase Y (CPY*) was found to be a luminal ER substrate and has served as a useful marker to help identify modifiers of the ERAD pathway. Due to its ease of genetic manipulation and the ability to conduct a genome wide screen for modifiers of molecular pathways, C. elegans has become one of the preferred metazoans for studying cell biological processes, such as ERAD. However, a marker of ERAD activity comparable to CPY* has not been developed for this model system. We describe a mutant of pro-cathepsin L fused to YFP that no longer targets to the lysosome, but is efficiently eliminated by the ERAD pathway. Using this mutant pro-cathepsin L, we found that components of the mammalian ERAD system that participate in the degradation of ER luminal substrates were conserved in C. elegans. This transgenic line will facilitate high-throughput genetic or pharmacological screens for ERAD modifiers using widefield epifluorescence microscopy. Public Library of Science 2012-07-02 /pmc/articles/PMC3388072/ /pubmed/22768338 http://dx.doi.org/10.1371/journal.pone.0040145 Text en Miedel et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Miedel, Mark T.
Graf, Nathan J.
Stephen, Kate E.
Long, Olivia S.
Pak, Stephen C.
Perlmutter, David H.
Silverman, Gary A.
Luke, Cliff J.
A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans
title A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans
title_full A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans
title_fullStr A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans
title_full_unstemmed A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans
title_short A Pro-Cathepsin L Mutant Is a Luminal Substrate for Endoplasmic-Reticulum-Associated Degradation in C. elegans
title_sort pro-cathepsin l mutant is a luminal substrate for endoplasmic-reticulum-associated degradation in c. elegans
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3388072/
https://www.ncbi.nlm.nih.gov/pubmed/22768338
http://dx.doi.org/10.1371/journal.pone.0040145
work_keys_str_mv AT miedelmarkt aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT grafnathanj aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT stephenkatee aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT longolivias aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT pakstephenc aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT perlmutterdavidh aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT silvermangarya aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT lukecliffj aprocathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT miedelmarkt procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT grafnathanj procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT stephenkatee procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT longolivias procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT pakstephenc procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT perlmutterdavidh procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT silvermangarya procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans
AT lukecliffj procathepsinlmutantisaluminalsubstrateforendoplasmicreticulumassociateddegradationincelegans