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High resolution imaging reveals heterogeneity in chromatin states between cells that is not inherited through cell division

View Article: PubMed Central - PubMed

ABSTRACT

Background: Genomes of eukaryotes exist as chromatin, and it is known that different chromatin states can influence gene regulation. Chromatin is not a static structure, but is known to be dynamic and vary between cells. In order to monitor the organisation of chromatin in live cells we have engineered fluorescent fusion proteins which recognize specific operator sequences to tag pairs of syntenic gene loci. The separation of these loci was then tracked in three dimensions over time using fluorescence microscopy.

Results: We established a work flow for measuring the distance between two fluorescently tagged, syntenic gene loci with a mean measurement error of 63 nm. In general, physical separation was observed to increase with increasing genomic separations. However, the extent to which chromatin is compressed varies for different genomic regions. No correlation was observed between compaction and the distribution of chromatin markers from genomic datasets or with contacts identified using capture based approaches. Variation in spatial separation was also observed within cells over time and between cells. Differences in the conformation of individual loci can persist for minutes in individual cells. Separation of reporter loci was found to be similar in related and unrelated daughter cell pairs.

Conclusions: The directly observed physical separation of reporter loci in live cells is highly dynamic both over time and from cell to cell. However, consistent differences in separation are observed over some chromosomal regions that do not correlate with factors known to influence chromatin states. We conclude that as yet unidentified parameters influence chromatin configuration. We also find that while heterogeneity in chromatin states can be maintained for minutes between cells, it is not inherited through cell division. This may contribute to cell-to-cell transcriptional heterogeneity.

Electronic supplementary material: The online version of this article (doi:10.1186/s12860-016-0111-y) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

Relationship between genomic separation and physical distance. a Frequency distributions of spot distances for the genomic separations indicated. Bin sizes are based on mean measurement error (63 nm). b Box plots representing spot distances for measurements from each strain. c Compaction ratio defined as 0.34α/d, where α is the genomic separation (in base pairs) and d is the measured distance (in nm)
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Fig2: Relationship between genomic separation and physical distance. a Frequency distributions of spot distances for the genomic separations indicated. Bin sizes are based on mean measurement error (63 nm). b Box plots representing spot distances for measurements from each strain. c Compaction ratio defined as 0.34α/d, where α is the genomic separation (in base pairs) and d is the measured distance (in nm)

Mentions: Using the workflow described above it was possible to measure the distance between two fluorescently tagged loci over time in several strains. Spot distance behaviours from all videos are presented in Additional file 1: Figure S2. Distance measurements for strains with varying genomic separations in G1 of the cell cycle are presented as histograms in Fig. 2a. When the distributions obtained from all strains are plotted in boxplot format, it is apparent that for the longer genomic separations there is a progressive but non-linear increase in the physical distance (Fig. 2b), similar to that previously reported [54].Fig. 2


High resolution imaging reveals heterogeneity in chromatin states between cells that is not inherited through cell division
Relationship between genomic separation and physical distance. a Frequency distributions of spot distances for the genomic separations indicated. Bin sizes are based on mean measurement error (63 nm). b Box plots representing spot distances for measurements from each strain. c Compaction ratio defined as 0.34α/d, where α is the genomic separation (in base pairs) and d is the measured distance (in nm)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5016949&req=5

Fig2: Relationship between genomic separation and physical distance. a Frequency distributions of spot distances for the genomic separations indicated. Bin sizes are based on mean measurement error (63 nm). b Box plots representing spot distances for measurements from each strain. c Compaction ratio defined as 0.34α/d, where α is the genomic separation (in base pairs) and d is the measured distance (in nm)
Mentions: Using the workflow described above it was possible to measure the distance between two fluorescently tagged loci over time in several strains. Spot distance behaviours from all videos are presented in Additional file 1: Figure S2. Distance measurements for strains with varying genomic separations in G1 of the cell cycle are presented as histograms in Fig. 2a. When the distributions obtained from all strains are plotted in boxplot format, it is apparent that for the longer genomic separations there is a progressive but non-linear increase in the physical distance (Fig. 2b), similar to that previously reported [54].Fig. 2

View Article: PubMed Central - PubMed

ABSTRACT

Background: Genomes of eukaryotes exist as chromatin, and it is known that different chromatin states can influence gene regulation. Chromatin is not a static structure, but is known to be dynamic and vary between cells. In order to monitor the organisation of chromatin in live cells we have engineered fluorescent fusion proteins which recognize specific operator sequences to tag pairs of syntenic gene loci. The separation of these loci was then tracked in three dimensions over time using fluorescence microscopy.

Results: We established a work flow for measuring the distance between two fluorescently tagged, syntenic gene loci with a mean measurement error of 63 nm. In general, physical separation was observed to increase with increasing genomic separations. However, the extent to which chromatin is compressed varies for different genomic regions. No correlation was observed between compaction and the distribution of chromatin markers from genomic datasets or with contacts identified using capture based approaches. Variation in spatial separation was also observed within cells over time and between cells. Differences in the conformation of individual loci can persist for minutes in individual cells. Separation of reporter loci was found to be similar in related and unrelated daughter cell pairs.

Conclusions: The directly observed physical separation of reporter loci in live cells is highly dynamic both over time and from cell to cell. However, consistent differences in separation are observed over some chromosomal regions that do not correlate with factors known to influence chromatin states. We conclude that as yet unidentified parameters influence chromatin configuration. We also find that while heterogeneity in chromatin states can be maintained for minutes between cells, it is not inherited through cell division. This may contribute to cell-to-cell transcriptional heterogeneity.

Electronic supplementary material: The online version of this article (doi:10.1186/s12860-016-0111-y) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus