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A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila
Single-molecule localization microscopy (SMLM) can provide nanoscale resolution in thin samples but has rarely been applied to tissues because of high background from out-of-focus emitters and optical aberrations. Here, we describe a line scanning microscope that provides optical sectioning for SMLM...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The Company of Biologists Ltd
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8729783/ https://www.ncbi.nlm.nih.gov/pubmed/34806753 http://dx.doi.org/10.1242/jcs.259570 |
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author | Cheng, Jinmei Allgeyer, Edward S. Richens, Jennifer H. Dzafic, Edo Palandri, Amandine Lewków, Bohdan Sirinakis, George St Johnston, Daniel |
author_facet | Cheng, Jinmei Allgeyer, Edward S. Richens, Jennifer H. Dzafic, Edo Palandri, Amandine Lewków, Bohdan Sirinakis, George St Johnston, Daniel |
author_sort | Cheng, Jinmei |
collection | PubMed |
description | Single-molecule localization microscopy (SMLM) can provide nanoscale resolution in thin samples but has rarely been applied to tissues because of high background from out-of-focus emitters and optical aberrations. Here, we describe a line scanning microscope that provides optical sectioning for SMLM in tissues. Imaging endogenously-tagged nucleoporins and F-actin on this system using DNA- and peptide-point accumulation for imaging in nanoscale topography (PAINT) routinely gives 30 nm resolution or better at depths greater than 20 µm. This revealed that the nuclear pores are nonrandomly distributed in most Drosophila tissues, in contrast to what is seen in cultured cells. Lamin Dm(0) shows a complementary localization to the nuclear pores, suggesting that it corrals the pores. Furthermore, ectopic expression of the tissue-specific Lamin C causes the nuclear pores to distribute more randomly, whereas lamin C mutants enhance nuclear pore clustering, particularly in muscle nuclei. Given that nucleoporins interact with specific chromatin domains, nuclear pore clustering could regulate local chromatin organization and contribute to the disease phenotypes caused by human lamin A/C laminopathies. |
format | Online Article Text |
id | pubmed-8729783 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | The Company of Biologists Ltd |
record_format | MEDLINE/PubMed |
spelling | pubmed-87297832022-01-12 A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila Cheng, Jinmei Allgeyer, Edward S. Richens, Jennifer H. Dzafic, Edo Palandri, Amandine Lewków, Bohdan Sirinakis, George St Johnston, Daniel J Cell Sci Research Article Single-molecule localization microscopy (SMLM) can provide nanoscale resolution in thin samples but has rarely been applied to tissues because of high background from out-of-focus emitters and optical aberrations. Here, we describe a line scanning microscope that provides optical sectioning for SMLM in tissues. Imaging endogenously-tagged nucleoporins and F-actin on this system using DNA- and peptide-point accumulation for imaging in nanoscale topography (PAINT) routinely gives 30 nm resolution or better at depths greater than 20 µm. This revealed that the nuclear pores are nonrandomly distributed in most Drosophila tissues, in contrast to what is seen in cultured cells. Lamin Dm(0) shows a complementary localization to the nuclear pores, suggesting that it corrals the pores. Furthermore, ectopic expression of the tissue-specific Lamin C causes the nuclear pores to distribute more randomly, whereas lamin C mutants enhance nuclear pore clustering, particularly in muscle nuclei. Given that nucleoporins interact with specific chromatin domains, nuclear pore clustering could regulate local chromatin organization and contribute to the disease phenotypes caused by human lamin A/C laminopathies. The Company of Biologists Ltd 2021-12-16 /pmc/articles/PMC8729783/ /pubmed/34806753 http://dx.doi.org/10.1242/jcs.259570 Text en © 2021. Published by The Company of Biologists Ltd https://creativecommons.org/licenses/by/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed. |
spellingShingle | Research Article Cheng, Jinmei Allgeyer, Edward S. Richens, Jennifer H. Dzafic, Edo Palandri, Amandine Lewków, Bohdan Sirinakis, George St Johnston, Daniel A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila |
title | A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila |
title_full | A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila |
title_fullStr | A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila |
title_full_unstemmed | A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila |
title_short | A single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in Drosophila |
title_sort | single-molecule localization microscopy method for tissues reveals nonrandom nuclear pore distribution in drosophila |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8729783/ https://www.ncbi.nlm.nih.gov/pubmed/34806753 http://dx.doi.org/10.1242/jcs.259570 |
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