Live imaging reveals chromatin compaction transitions and dynamic transcriptional bursting during stem cell differentiation in vivo
- Department of Genetics, Yale University School of Medicine, United States
- Institute for Quantitative Health Science & Engineering (IQ), Michigan State University, United States
- Division of Dermatology, Department of Medicine, College of Human Medicine, Michigan State University, United States
- Department of Pharmacology and Toxicology, College of Human Medicine, Michigan State University, United States
- Department of Biostatistics, Yale University School of Public Health, United States
- Deparment of Cell Biology, Yale University School of Medicine, United States
- Department of Biochemistry and Rosalind & Morris Goodman Cancer Institute, McGill University, Canada
- Departments of Cell Biology and Dermatology, Yale Stem Cell Center, Yale Cancer Center, Yale University School of Medicine, United States
Figures
Figure 1 with 2 supplements
Chromatin compaction state is heterogeneous and independent of interphase cell cycle.
(A) Representative XY view of the basal stem cell layer showing the Kertain14-driven Histone2B-GFP allele in a live mouse. (B) A representative chromatin compaction profile of a single basal stem …
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Figure 1—source data 1
Heterogeneous chromatin compaction independent of interphase cell cycle status.
- https://cdn.elifesciences.org/articles/83444/elife-83444-fig1-data1-v2.xlsx
Figure 1—figure supplement 1
Chromatin compaction state analysis.
(A) XZ schematic of the mouse epidermis. The basal stem cell layer (bottom) contains actively cycling cells as well as cells that have committed to differentiation and are actively leaving the …
Figure 1—figure supplement 2
Organization of H2B-GFP allele.
(A) Representative crops of fixed tissue, single channel stainings of the basal stem cell layer. Genetically encoded K14H2B-GFP signal is shown in green, and immunofluorescent stainings in magenta. …
Figure 2 with 1 supplement
Chromatin compaction state changes through differentiation state.
(A) XZ schematic of the epidermis. The basal stem cell layer is shown with black nuclei and the differentiated (spinous) layer is shown apical with red nuclei. (B) Representative crops of individual …
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Figure 2—source data 1
Chromatin compaction state changes with differentiation status.
- https://cdn.elifesciences.org/articles/83444/elife-83444-fig2-data1-v2.xlsx
Figure 2—figure supplement 1
Genetic ablation of SRF alle in the basal stem cell layer.
(A) Schematic of the inducible genetic system used in Figure 2 to recombine and knock down Srf. (B) Representative crops from the basal stem cell and differentiated layers on days 0 and 6 in …
Figure 3 with 1 supplement
Chromatin compaction is stable over hours and progressively changes over days.
(A) Time lapse imaging data of a single nucleus crop at 0, 1.5, and 3 hr time points. H2B-GFP fluorescent signal is shown in white. Three, high-intensity chromocenters were chosen and pseudo-colored …
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Figure 3—source data 1
Chromatin compaction state changes slowly over days.
- https://cdn.elifesciences.org/articles/83444/elife-83444-fig3-data1-v2.xlsx
Figure 3—figure supplement 1
Single-cell chromatin compaction dynamics over hours.
(A) Individual nucleus chromatin compaction profiles from 3 basal stem cell and 3 delaminating/spinning cells at 0-, 1.5-, and 3 hr time points showing little change over this time scale. (B) …
Figure 4
Chromatin compaction changes precede differentiation.
(A) XY schematic of genetic system (K14H2B-GFP; K10rtTA; tetO-Cre; R26LSL-tdTomato) allowing visualization of actively differentiating cells still within the basal stem cell layer (expressing …
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Figure 4—source data 1
Chromatin compaction state begins remodeling upstream of basal delamination.
- https://cdn.elifesciences.org/articles/83444/elife-83444-fig4-data1-v2.xlsx
Figure 5 with 1 supplement
in vivo transcription of Keratin-10 precedes genome architecture changes through differentiation.
(A) Visual schematic of the MCP/MS2 system allowing visualization of a targeted gene under endogenous regulation. 24X MS2 repeats were knocked into the 3′UTR of the Keratin-10 locus and chromatin …
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Figure 5—source data 1
Transcription of differentiation gene is highly dynamic and precedes significant chromatin compaction changes.
- https://cdn.elifesciences.org/articles/83444/elife-83444-fig5-data1-v2.xlsx
Figure 5—figure supplement 1
Imaging differentiation-associated cell identity.
(A) Representative crops of the differentiated spinous layer (left) and basal stem cell layer (right), as well as magnified inset showing active site of Keratin-10 transcription, and nascent …
Figure 6
Chromatin architecture remodeling through epidermal differentiation.
Epidermal stem cells undergo incremental changes toward differentiation over 3–4 days (top). During this process, differentiation-committed cells within the basal stem cell layer begin expressing Ker…
Author response image 1
K10MS2/MCP punctum do not colocalize with the lowest H2BGFP fluorescent regions.
Videos
Video 1
Chromatin compaction.
A single basal stem cell layer nucleus crop visualizing H2B-GFP intensity. The video first scans through the greyscale, intensity image (white), then through the FIRE LUT intensities shown in Figure …
Video 2
Spinning chromatin.
An XY field-of-view of the upper, basal stem cell layer in which chromatin (H2B-GFP) can be observed to spin. Timelapse is 3 hours long and looped 3 times.
