Live imaging reveals chromatin compaction transitions and dynamic transcriptional bursting during stem cell differentiation in vivo
Abstract
Stem cell differentiation requires dramatic changes in gene expression and global remodeling of chromatin architecture. How and when chromatin remodels relative to the transcriptional, behavioral, and morphological changes during differentiation remain unclear, particularly in an intact tissue context. Here, we develop a quantitative pipeline which leverages fluorescently-tagged histones and longitudinal imaging to track large-scale chromatin compaction changes within individual cells in a live mouse. Applying this pipeline to epidermal stem cells, we reveal that cell-to-cell chromatin compaction heterogeneity within the stem cell compartment emerges independent of cell cycle status, and instead is reflective of differentiation status. Chromatin compaction state gradually transitions over days as differentiating cells exit the stem cell compartment. Moreover, establishing live imaging of Keratin-10 (K10) nascent RNA, which marks the onset of stem cell differentiation, we find that Keratin-10 transcription is highly dynamic and largely precedes the global chromatin compaction changes associated with differentiation. Together, these analyses reveal that stem cell differentiation involves dynamic transcriptional states and gradual chromatin rearrangement.
Data availability
All coding scripts and source datasheets for figure quantifications are made accessible through the Dryad data repository: https://doi.org/10.5061/dryad.5hqbzkh94. Representative raw imaging data are accessible through the same link and full datasets available upon request with no restrictions (due to size) by contacting VG. Source datasheets are included in supplemental information.
Article and author information
Author details
Funding
National Institute of Arthritis and Musculoskeletal and Skin Diseases (1R01AR063663-01)
- Valentina Greco
National Institute of Arthritis and Musculoskeletal and Skin Diseases (1R01AR067755-01A1)
- Valentina Greco
National Institute on Aging (1DP1AG066590-01)
- Valentina Greco
National Institute of Arthritis and Musculoskeletal and Skin Diseases (R01AR072668)
- Valentina Greco
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: All procedures involving animal subjects were performed under the approval of the Institutional Animal Care and Use Committee (IACUC) of the Yale School of Medicine (Protocol #2021-11303). All live imaging was performed under 1-2% isoflurane, and ever effort was made to minimize suffering.
Copyright
© 2023, May et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 4,585
- views
-
- 473
- downloads
-
- 6
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Developmental Biology
- Stem Cells and Regenerative Medicine
Amniogenesis, a process critical for continuation of healthy pregnancy, is triggered in a collection of pluripotent epiblast cells as the human embryo implants. Previous studies have established that bone morphogenetic protein (BMP) signaling is a major driver of this lineage specifying process, but the downstream BMP-dependent transcriptional networks that lead to successful amniogenesis remain to be identified. This is, in part, due to the current lack of a robust and reproducible model system that enables mechanistic investigations exclusively into amniogenesis. Here, we developed an improved model of early amnion specification, using a human pluripotent stem cell-based platform in which the activation of BMP signaling is controlled and synchronous. Uniform amniogenesis is seen within 48 hr after BMP activation, and the resulting cells share transcriptomic characteristics with amnion cells of a gastrulating human embryo. Using detailed time-course transcriptomic analyses, we established a previously uncharacterized BMP-dependent amniotic transcriptional cascade, and identified markers that represent five distinct stages of amnion fate specification; the expression of selected markers was validated in early post-implantation macaque embryos. Moreover, a cohort of factors that could potentially control specific stages of amniogenesis was identified, including the transcription factor TFAP2A. Functionally, we determined that, once amniogenesis is triggered by the BMP pathway, TFAP2A controls the progression of amniogenesis. This work presents a temporally resolved transcriptomic resource for several previously uncharacterized amniogenesis states and demonstrates a critical intermediate role for TFAP2A during amnion fate specification.
-
- Stem Cells and Regenerative Medicine
A new mathematical model can estimate the number of precursor cells that contribute to regenerating blood cells in mice.