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Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system
Functional integrity of eukaryotic organelles relies on direct physical contacts between distinct organelles. However, the entity of organelle-tethering factors is not well understood due to lack of means to analyze inter-organelle interactions in living cells. Here we evaluate the split-GFP system...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5906596/ https://www.ncbi.nlm.nih.gov/pubmed/29670150 http://dx.doi.org/10.1038/s41598-018-24466-0 |
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author | Kakimoto, Yuriko Tashiro, Shinya Kojima, Rieko Morozumi, Yuki Endo, Toshiya Tamura, Yasushi |
author_facet | Kakimoto, Yuriko Tashiro, Shinya Kojima, Rieko Morozumi, Yuki Endo, Toshiya Tamura, Yasushi |
author_sort | Kakimoto, Yuriko |
collection | PubMed |
description | Functional integrity of eukaryotic organelles relies on direct physical contacts between distinct organelles. However, the entity of organelle-tethering factors is not well understood due to lack of means to analyze inter-organelle interactions in living cells. Here we evaluate the split-GFP system for visualizing organelle contact sites in vivo and show its advantages and disadvantages. We observed punctate GFP signals from the split-GFP fragments targeted to any pairs of organelles among the ER, mitochondria, peroxisomes, vacuole and lipid droplets in yeast cells, which suggests that these organelles form contact sites with multiple organelles simultaneously although it is difficult to rule out the possibilities that these organelle contacts sites are artificially formed by the irreversible associations of the split-GFP probes. Importantly, split-GFP signals in the overlapped regions of the ER and mitochondria were mainly co-localized with ERMES, an authentic ER-mitochondria tethering structure, suggesting that split-GFP assembly depends on the preexisting inter-organelle contact sites. We also confirmed that the split-GFP system can be applied to detection of the ER-mitochondria contact sites in HeLa cells. We thus propose that the split-GFP system is a potential tool to observe and analyze inter-organelle contact sites in living yeast and mammalian cells. |
format | Online Article Text |
id | pubmed-5906596 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-59065962018-04-30 Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system Kakimoto, Yuriko Tashiro, Shinya Kojima, Rieko Morozumi, Yuki Endo, Toshiya Tamura, Yasushi Sci Rep Article Functional integrity of eukaryotic organelles relies on direct physical contacts between distinct organelles. However, the entity of organelle-tethering factors is not well understood due to lack of means to analyze inter-organelle interactions in living cells. Here we evaluate the split-GFP system for visualizing organelle contact sites in vivo and show its advantages and disadvantages. We observed punctate GFP signals from the split-GFP fragments targeted to any pairs of organelles among the ER, mitochondria, peroxisomes, vacuole and lipid droplets in yeast cells, which suggests that these organelles form contact sites with multiple organelles simultaneously although it is difficult to rule out the possibilities that these organelle contacts sites are artificially formed by the irreversible associations of the split-GFP probes. Importantly, split-GFP signals in the overlapped regions of the ER and mitochondria were mainly co-localized with ERMES, an authentic ER-mitochondria tethering structure, suggesting that split-GFP assembly depends on the preexisting inter-organelle contact sites. We also confirmed that the split-GFP system can be applied to detection of the ER-mitochondria contact sites in HeLa cells. We thus propose that the split-GFP system is a potential tool to observe and analyze inter-organelle contact sites in living yeast and mammalian cells. Nature Publishing Group UK 2018-04-18 /pmc/articles/PMC5906596/ /pubmed/29670150 http://dx.doi.org/10.1038/s41598-018-24466-0 Text en © The Author(s) 2018 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Kakimoto, Yuriko Tashiro, Shinya Kojima, Rieko Morozumi, Yuki Endo, Toshiya Tamura, Yasushi Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system |
title | Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system |
title_full | Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system |
title_fullStr | Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system |
title_full_unstemmed | Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system |
title_short | Visualizing multiple inter-organelle contact sites using the organelle-targeted split-GFP system |
title_sort | visualizing multiple inter-organelle contact sites using the organelle-targeted split-gfp system |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5906596/ https://www.ncbi.nlm.nih.gov/pubmed/29670150 http://dx.doi.org/10.1038/s41598-018-24466-0 |
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