Cargando…
Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy
In eukaryotic cells, highly condensed inactive/silenced chromatin has long been called “heterochromatin.” However, recent research suggests that such regions are in fact not fully transcriptionally silent and that there exists only a moderate access barrier to heterochromatin. To further investigate...
Autores principales: | , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The American Society for Cell Biology
2017
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5687035/ https://www.ncbi.nlm.nih.gov/pubmed/28835378 http://dx.doi.org/10.1091/mbc.E17-06-0359 |
_version_ | 1783278895547547648 |
---|---|
author | Imai, Ryosuke Nozaki, Tadasu Tani, Tomomi Kaizu, Kazunari Hibino, Kayo Ide, Satoru Tamura, Sachiko Takahashi, Koichi Shribak, Michael Maeshima, Kazuhiro |
author_facet | Imai, Ryosuke Nozaki, Tadasu Tani, Tomomi Kaizu, Kazunari Hibino, Kayo Ide, Satoru Tamura, Sachiko Takahashi, Koichi Shribak, Michael Maeshima, Kazuhiro |
author_sort | Imai, Ryosuke |
collection | PubMed |
description | In eukaryotic cells, highly condensed inactive/silenced chromatin has long been called “heterochromatin.” However, recent research suggests that such regions are in fact not fully transcriptionally silent and that there exists only a moderate access barrier to heterochromatin. To further investigate this issue, it is critical to elucidate the physical properties of heterochromatin such as its total density in live cells. Here, using orientation-independent differential interference contrast (OI-DIC) microscopy, which is capable of mapping optical path differences, we investigated the density of the total materials in pericentric foci, a representative heterochromatin model, in live mouse NIH3T3 cells. We demonstrated that the total density of heterochromatin (208 mg/ml) was only 1.53-fold higher than that of the surrounding euchromatic regions (136 mg/ml) while the DNA density of heterochromatin was 5.5- to 7.5-fold higher. We observed similar minor differences in density in typical facultative heterochromatin, the inactive human X chromosomes. This surprisingly small difference may be due to that nonnucleosomal materials (proteins/RNAs) (∼120 mg/ml) are dominant in both chromatin regions. Monte Carlo simulation suggested that nonnucleosomal materials contribute to creating a moderate access barrier to heterochromatin, allowing minimal protein access to functional regions. Our OI-DIC imaging offers new insight into the live cellular environments. |
format | Online Article Text |
id | pubmed-5687035 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | The American Society for Cell Biology |
record_format | MEDLINE/PubMed |
spelling | pubmed-56870352018-01-22 Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy Imai, Ryosuke Nozaki, Tadasu Tani, Tomomi Kaizu, Kazunari Hibino, Kayo Ide, Satoru Tamura, Sachiko Takahashi, Koichi Shribak, Michael Maeshima, Kazuhiro Mol Biol Cell Articles In eukaryotic cells, highly condensed inactive/silenced chromatin has long been called “heterochromatin.” However, recent research suggests that such regions are in fact not fully transcriptionally silent and that there exists only a moderate access barrier to heterochromatin. To further investigate this issue, it is critical to elucidate the physical properties of heterochromatin such as its total density in live cells. Here, using orientation-independent differential interference contrast (OI-DIC) microscopy, which is capable of mapping optical path differences, we investigated the density of the total materials in pericentric foci, a representative heterochromatin model, in live mouse NIH3T3 cells. We demonstrated that the total density of heterochromatin (208 mg/ml) was only 1.53-fold higher than that of the surrounding euchromatic regions (136 mg/ml) while the DNA density of heterochromatin was 5.5- to 7.5-fold higher. We observed similar minor differences in density in typical facultative heterochromatin, the inactive human X chromosomes. This surprisingly small difference may be due to that nonnucleosomal materials (proteins/RNAs) (∼120 mg/ml) are dominant in both chromatin regions. Monte Carlo simulation suggested that nonnucleosomal materials contribute to creating a moderate access barrier to heterochromatin, allowing minimal protein access to functional regions. Our OI-DIC imaging offers new insight into the live cellular environments. The American Society for Cell Biology 2017-11-07 /pmc/articles/PMC5687035/ /pubmed/28835378 http://dx.doi.org/10.1091/mbc.E17-06-0359 Text en © 2017 Imai et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology. |
spellingShingle | Articles Imai, Ryosuke Nozaki, Tadasu Tani, Tomomi Kaizu, Kazunari Hibino, Kayo Ide, Satoru Tamura, Sachiko Takahashi, Koichi Shribak, Michael Maeshima, Kazuhiro Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy |
title | Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy |
title_full | Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy |
title_fullStr | Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy |
title_full_unstemmed | Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy |
title_short | Density imaging of heterochromatin in live cells using orientation-independent-DIC microscopy |
title_sort | density imaging of heterochromatin in live cells using orientation-independent-dic microscopy |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5687035/ https://www.ncbi.nlm.nih.gov/pubmed/28835378 http://dx.doi.org/10.1091/mbc.E17-06-0359 |
work_keys_str_mv | AT imairyosuke densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT nozakitadasu densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT tanitomomi densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT kaizukazunari densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT hibinokayo densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT idesatoru densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT tamurasachiko densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT takahashikoichi densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT shribakmichael densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy AT maeshimakazuhiro densityimagingofheterochromatininlivecellsusingorientationindependentdicmicroscopy |