The life cycle of microorganisms is associated with dynamic metabolic transitions and complex cellular responses. In yeast, how metabolic signals control the progressive choreography of structural reorganizations observed in quiescent cells during a natural life cycle remains unclear. We have developed an integrated microfluidic device to address this question, enabling continuous single-cell tracking in a batch culture experiencing unperturbed nutrient exhaustion to unravel the coordination between metabolic and structural transitions within cells. Our technique reveals an abrupt fate divergence in the population, whereby a fraction of cells is unable to transition to respiratory metabolism and undergoes a reversible entry into a quiescence-like state leading to premature cell death. Further observations reveal that non-monotonous internal pH fluctuations in respiration-competent cells orchestrate the successive waves of protein super-assemblies formation that accompany the entry into a bona fide quiescent state. This ultimately leads to an abrupt cytosolic glass transition that occurs stochastically long after proliferation cessation. This new experimental framework provides a unique way to track single-cell fate dynamics over a long timescale in a population of cells that continuously modify their ecological niche.
The CAD file used to generate the microfluidic device is available on a github repository. The source data used to make the panels (excluding raw image files) are included for each figure. Due to size constraints representative raw image data for Figure 1 is available at Zenodo (https://doi.org/10.5281/zenodo.5592983) and the remaining raw image data, including files for Figures 2 and 3, are available on request from the corresponding author.
Dataset pHluorin cells experiencing entry into quiescenceZenodo, doi:10.5281/zenodo.5592983.
- Basile Jacquel
- Théo Aspert
- Gilles Charvin
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Bavesh D Kana, University of the Witwatersrand, South Africa
© 2021, Jacquel 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.
Neonatal cerebral hypoxia-ischemia (HI) is the leading cause of death and disability in newborns with the only current treatment being hypothermia. An increased understanding of the pathways that facilitate tissue repair after HI may aid the development of better treatments. Here, we study the role of lactate receptor HCAR1 in tissue repair after neonatal HI in mice. We show that HCAR1 knockout mice have reduced tissue regeneration compared with wildtype mice. Furthermore, proliferation of neural progenitor cells and glial cells, as well as microglial activation was impaired. Transcriptome analysis showed a strong transcriptional response to HI in the subventricular zone of wildtype mice involving about 7300 genes. In contrast, the HCAR1 knockout mice showed a modest response, involving about 750 genes. Notably, fundamental processes in tissue repair such as cell cycle and innate immunity were dysregulated in HCAR1 knockout. Our data suggest that HCAR1 is a key transcriptional regulator of pathways that promote tissue regeneration after HI.
Btg3-associated nuclear protein (Banp) was originally identified as a nuclear matrix-associated region (MAR)-binding protein and it functions as a tumor suppressor. At the molecular level, Banp regulates transcription of metabolic genes via a CGCG-containing motif called the Banp motif. However, its physiological roles in embryonic development are unknown. Here, we report that Banp is indispensable for the DNA damage response and chromosome segregation during mitosis. Zebrafish banp mutants show mitotic cell accumulation and apoptosis in developing retina. We found that DNA replication stress and tp53-dependent DNA damage responses were activated to induce apoptosis in banp mutants, suggesting that Banp is required for regulation of DNA replication and DNA damage repair. Furthermore, consistent with mitotic cell accumulation, chromosome segregation was not smoothly processed from prometaphase to anaphase in banp morphants, leading to a prolonged M-phase. Our RNA- and ATAC-sequencing identified 31 candidates for direct Banp target genes that carry the Banp motif. Interestingly, a DNA replication fork regulator, wrnip1, and two chromosome segregation regulators, cenpt and ncapg, are included in this list. Thus, Banp directly regulates transcription of wrnip1 for recovery from DNA replication stress, and cenpt and ncapg for chromosome segregation during mitosis. Our findings provide the first in vivo evidence that Banp is required for cell-cycle progression and cell survival by regulating DNA damage responses and chromosome segregation during mitosis.