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Active Nuclear Import of Membrane Proteins Revisited
It is poorly understood how membrane proteins destined for the inner nuclear membrane pass the crowded environment of the Nuclear Pore Complex (NPC). For the Saccharomyces cerevisiae proteins Src1/Heh1 and Heh2, a transport mechanism was proposed where the transmembrane domains diffuse through the m...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4695851/ https://www.ncbi.nlm.nih.gov/pubmed/26473931 http://dx.doi.org/10.3390/cells4040653 |
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author | Laba, Justyna K. Steen, Anton Popken, Petra Chernova, Alina Poolman, Bert Veenhoff, Liesbeth M. |
author_facet | Laba, Justyna K. Steen, Anton Popken, Petra Chernova, Alina Poolman, Bert Veenhoff, Liesbeth M. |
author_sort | Laba, Justyna K. |
collection | PubMed |
description | It is poorly understood how membrane proteins destined for the inner nuclear membrane pass the crowded environment of the Nuclear Pore Complex (NPC). For the Saccharomyces cerevisiae proteins Src1/Heh1 and Heh2, a transport mechanism was proposed where the transmembrane domains diffuse through the membrane while the extralumenal domains encoding a nuclear localization signal (NLS) and intrinsically disordered linker (L) are accompanied by transport factors and travel through the NPC. Here, we validate the proposed mechanism and explore and discuss alternative interpretations of the data. First, to disprove an interpretation where the membrane proteins become membrane embedded only after nuclear import, we present biochemical and localization data to support that the previously used, as well as newly designed reporter proteins are membrane-embedded irrespective of the presence of the sorting signals, the specific transmembrane domain (multipass or tail anchored), independent of GET, and also under conditions that the proteins are trapped in the NPC. Second, using the recently established size limit for passive diffusion of membrane proteins in yeast, and using an improved assay, we confirm active import of polytopic membrane protein with extralumenal soluble domains larger than those that can pass by diffusion on similar timescales. This reinforces that NLS-L dependent active transport is distinct from passive diffusion. Thirdly, we revisit the proposed route through the center of the NPC and conclude that the previously used trapping assay is, unfortunately, poorly suited to address the route through the NPC, and the route thus remains unresolved. Apart from the uncertainty about the route through the NPC, the data confirm active, transport factor dependent, nuclear transport of membrane-embedded mono- and polytopic membrane proteins in baker’s yeast. |
format | Online Article Text |
id | pubmed-4695851 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-46958512016-01-19 Active Nuclear Import of Membrane Proteins Revisited Laba, Justyna K. Steen, Anton Popken, Petra Chernova, Alina Poolman, Bert Veenhoff, Liesbeth M. Cells Article It is poorly understood how membrane proteins destined for the inner nuclear membrane pass the crowded environment of the Nuclear Pore Complex (NPC). For the Saccharomyces cerevisiae proteins Src1/Heh1 and Heh2, a transport mechanism was proposed where the transmembrane domains diffuse through the membrane while the extralumenal domains encoding a nuclear localization signal (NLS) and intrinsically disordered linker (L) are accompanied by transport factors and travel through the NPC. Here, we validate the proposed mechanism and explore and discuss alternative interpretations of the data. First, to disprove an interpretation where the membrane proteins become membrane embedded only after nuclear import, we present biochemical and localization data to support that the previously used, as well as newly designed reporter proteins are membrane-embedded irrespective of the presence of the sorting signals, the specific transmembrane domain (multipass or tail anchored), independent of GET, and also under conditions that the proteins are trapped in the NPC. Second, using the recently established size limit for passive diffusion of membrane proteins in yeast, and using an improved assay, we confirm active import of polytopic membrane protein with extralumenal soluble domains larger than those that can pass by diffusion on similar timescales. This reinforces that NLS-L dependent active transport is distinct from passive diffusion. Thirdly, we revisit the proposed route through the center of the NPC and conclude that the previously used trapping assay is, unfortunately, poorly suited to address the route through the NPC, and the route thus remains unresolved. Apart from the uncertainty about the route through the NPC, the data confirm active, transport factor dependent, nuclear transport of membrane-embedded mono- and polytopic membrane proteins in baker’s yeast. MDPI 2015-10-13 /pmc/articles/PMC4695851/ /pubmed/26473931 http://dx.doi.org/10.3390/cells4040653 Text en © 2015 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Laba, Justyna K. Steen, Anton Popken, Petra Chernova, Alina Poolman, Bert Veenhoff, Liesbeth M. Active Nuclear Import of Membrane Proteins Revisited |
title | Active Nuclear Import of Membrane Proteins Revisited |
title_full | Active Nuclear Import of Membrane Proteins Revisited |
title_fullStr | Active Nuclear Import of Membrane Proteins Revisited |
title_full_unstemmed | Active Nuclear Import of Membrane Proteins Revisited |
title_short | Active Nuclear Import of Membrane Proteins Revisited |
title_sort | active nuclear import of membrane proteins revisited |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4695851/ https://www.ncbi.nlm.nih.gov/pubmed/26473931 http://dx.doi.org/10.3390/cells4040653 |
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