Dataset Information

Dose-dependent spatiotemporal responses of mammalian cells to an alkylating agent.

ABSTRACT: Cultured cell populations are composed of heterogeneous cells, and previous single-cell lineage tracking analysis of individual HeLa cells provided empirical evidence for significant heterogeneity of the rate of cell proliferation and induction of cell death. Nevertheless, such cell lines have been used for investigations of cellular responses to various substances, resulting in incomplete characterizations. This problem caused by heterogeneity within cell lines could be overcome by investigating the spatiotemporal responses of individual cells to a substance. However, no approach to investigate the responses by analyzing spatiotemporal data is currently available. Thus, this study aimed to analyze the spatiotemporal responses of individual HeLa cells to cytotoxic, sub-cytotoxic, and non-cytotoxic doses of the well-characterized carcinogen, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Although cytotoxic doses of MNNG are known to induce cell death, the single-cell tracking approach revealed that cell death occurred following at least four different cellular events, suggesting that cell death is induced via multiple processes. We also found that HeLa cells exposed to a sub-cytotoxic dose of MNNG were in a state of equilibrium between cell proliferation and cell death, with cell death again induced through different processes. However, exposure of cells to a non-cytotoxic dose of MNNG promoted growth by reducing the cell doubling time, thus promoting the growth of a sub-population of cells. A single-cell lineage tracking approach could dissect processes leading to cell death in a spatiotemporal manner and the results suggest that spatiotemporal data obtained by tracking individual cells can be used as a new type of bioinformatics data resource that enables the examination of cellular responses to various external substances.

SUBMITTER: Rancourt A

PROVIDER: S-EPMC6440626 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Dose-dependent spatiotemporal responses of mammalian cells to an alkylating agent.

Rancourt Ann A Sato Sachiko S Satoh Masahiko S MS

PloS one 20190329 3

Cultured cell populations are composed of heterogeneous cells, and previous single-cell lineage tracking analysis of individual HeLa cells provided empirical evidence for significant heterogeneity of the rate of cell proliferation and induction of cell death. Nevertheless, such cell lines have been used for investigations of cellular responses to various substances, resulting in incomplete characterizations. This problem caused by heterogeneity within cell lines could be overcome by investigatin ...[more]

PMID: 30925183

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Project description:Wnt/β-catenin signaling is a highly conserved molecular pathway that plays a crucial role in stem/progenitor systems and cancer. β-catenin, the main Wnt pathway effector, has two pools within a cell: one for cell-cell adhesion at the membrane and the other for transcriptional functions in the nucleus. However, the mechanism by which β-catenin mediates both roles remain unclear. The tightly controlled, well characterized system of nephrogenesis is an ideal model to decouple the roles of β-catenin at the membrane and in the nucleus. In kidney development, a delicate balance of nephron progenitor cell self-renewal and differentiation is required for the mesenchymal to epithelial transition (MET) in nephrogenesis and is driven by Wnt/β-catenin signaling. Given an ability to isolate and manipulate large numbers of NPCs in tissue culture, we can dissect the dual nature of β-catenin as a transcriptional activator and component of a cell membrane complex in adhesion. We pioneered a method using CRISPR/Cas9 gene editing to rapidly remove β-catenin, Tcf/Lef factors and simultaneous cadherin genes in primary NPCs. We have characterized the effects of modulating β-catenin and integrated RNA-seq results from β-catenin’s removal with mouse ChIP-seq and mouse single cell RNA -seq data. Functional analysis of β-catenin removal provides strong evidence for β-catenin regulation of NPC proliferation, independent of a direct Lef/Tcf associated transcriptional program. Together these data suggest β-catenin mediates aggregation, the first step in MET, through β-catenin mediated cell adhesion complexes while simultaneous transcriptional activation within these structures initiates the nephrogenic program. The studies provide new insight into the direct transcriptional role of Lef/Tcf/β-catenin complexes associated with the initiation of a nephron forming program. Overall, this study enhances an understanding of the molecular mechanisms underlying kidney development and the dual nature of β-catenin stem/progenitor systems at large.

Dataset Information

Dose-dependent spatiotemporal responses of mammalian cells to an alkylating agent.

Publications

Dose-dependent spatiotemporal responses of mammalian cells to an alkylating agent.

Similar Datasets

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