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Dictyostelium discoideum: An Alternative Nonanimal Model for Developmental Toxicity Testing.

ABSTRACT: A critical aspect of toxicity evaluation is developmental and reproductive toxicity (DART) testing. Traditionally, DART testing has been conducted in vivo in mammalian model systems. New legislation aimed at reducing animal use and the prohibitive costs associated with DART testing, together with a need to understand the genetic pathways underlying developmental toxicity means there is a growing demand for alternative model systems for toxicity evaluation. Here we explore the potential of the eukaryotic social amoeba Dictyostelium discoideum, which is already widely used as a simple model system for cell and developmental biology, as a potential nonanimal model for DART testing. We developed assays for high-throughput screening of toxicity during D. discoideum growth and development. This allowed the toxicity of a broad range of test compounds to be characterized, which revealed that D. discoideum can broadly predict mammalian toxicity. In addition, we show that this system can be used to perform functional genomic screens to compare the molecular modes of action of different compounds. For example, genome-wide screens for mutations that affect lithium and valproic acid toxicity allowed common and unique biological targets and molecular processes mediating their toxicity to be identified. These studies illustrate that D. discoideum could represent a predictive nonanimal model for DART testing due to its amenability to high-throughput approaches and molecular genetic tractability.

SUBMITTER: Baines RP

PROVIDER: S-EPMC8538044 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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Dictyostelium discoideum: An Alternative Nonanimal Model for Developmental Toxicity Testing.

Baines Robert P RP Wolton Kathryn K Thompson Christopher R L CRL

Toxicological sciences : an official journal of the Society of Toxicology 20210901 2

A critical aspect of toxicity evaluation is developmental and reproductive toxicity (DART) testing. Traditionally, DART testing has been conducted in vivo in mammalian model systems. New legislation aimed at reducing animal use and the prohibitive costs associated with DART testing, together with a need to understand the genetic pathways underlying developmental toxicity means there is a growing demand for alternative model systems for toxicity evaluation. Here we explore the potential of the eu ...[more]

PMID: 34387693

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Project description:BackgroundDevelopment of the soil amoeba Dictyostelium discoideum is triggered by starvation. When placed on a solid substrate, the starving solitary amoebae cease growth, communicate via extracellular cAMP, aggregate by tens of thousands and develop into multicellular organisms. Early phases of the developmental program are often studied in cells starved in suspension while cAMP is provided exogenously. Previous studies revealed massive shifts in the transcriptome under both developmental conditions and a close relationship between gene expression and morphogenesis, but were limited by the sampling frequency and the resolution of the methods.ResultsHere, we combine the superior depth and specificity of RNA-seq-based analysis of mRNA abundance with high frequency sampling during filter development and cAMP pulsing in suspension. We found that the developmental transcriptome exhibits mostly gradual changes interspersed by a few instances of large shifts. For each time point we treated the entire transcriptome as single phenotype, and were able to characterize development as groups of similar time points separated by gaps. The grouped time points represented gradual changes in mRNA abundance, or molecular phenotype, and the gaps represented times during which many genes are differentially expressed rapidly, and thus the phenotype changes dramatically. Comparing developmental experiments revealed that gene expression in filter developed cells lagged behind those treated with exogenous cAMP in suspension. The high sampling frequency revealed many genes whose regulation is reproducibly more complex than indicated by previous studies. Gene Ontology enrichment analysis suggested that the transition to multicellularity coincided with rapid accumulation of transcripts associated with DNA processes and mitosis. Later development included the up-regulation of organic signaling molecules and co-factor biosynthesis. Our analysis also demonstrated a high level of synchrony among the developing structures throughout development.ConclusionsOur data describe D. discoideum development as a series of coordinated cellular and multicellular activities. Coordination occurred within fields of aggregating cells and among multicellular bodies, such as mounds or migratory slugs that experience both cell-cell contact and various soluble signaling regimes. These time courses, sampled at the highest temporal resolution to date in this system, provide a comprehensive resource for studies of developmental gene expression.

Project description:Development of the soil amoeba Dictyostelium discoideum is triggered by starvation. When placed on a solid substrate, the starving solitary amoebae cease growth, communicate via extracellular cAMP, aggregate by tens of thousands and develop into fully integrated multicellular organisms. Early phases of the developmental program are often studied in cells starved in suspension while cAMP is provided exogenously. Previous studies revealed massive shifts in the transcriptome under these developmental conditions and a close relationship between gene expression and morphogenesis, but were limited by the sampling frequency and the resolution of the methods. Here, we combine the superior depth and specificity of RNA-seq with high frequency sampling during filter development and cAMP pulsing in suspension. We found that the developmental transcriptome exhibits mostly gradual changes interspersed by a few instances of large changes. Considering the whole transcriptome as a single phenotype, we treated each time point as an independent entity and were able to characterize development as groups of similar time points separated by gaps. The grouped time points represent gradual changes in mRNA abundance and the gaps represent times during which many genes are differentially expressed rapidly. Comparing development on solid substrate to development in suspension revealed that gene expression in filter developed cells lagged behind cells treated with exogenous cAMP in suspension. The high sampling frequency revealed many genes whose regulation is reproducibly more complex than indicated by previous studies. Gene Ontology enrichment analysis showed that the transition to multicellularity at the mound stage coincided with rapid accumulation of transcripts associated with mitosis and DNA replicative processes. Later development included the up-regulation of organic signaling molecules and co-factor biosynthesis. We observed multiple instances of enrichment of oxidation-reduction and reactive oxygen related terms. Our analysis also demonstrated a high level of synchrony between the developing structures throughout development. Our data describe D. discoideum development as a series of coordinated cellular and multicellular activities. Coordination occurred within fields of aggregating cells and between multicellular bodies, such as mounds or migratory slugs that experience both cell-cell contact and various soluble signaling regimes. These time courses, sampled at the highest temporal resolution to date in this system, provide a comprehensive resource for future studies of developmental gene expression.

Dataset Information

Dictyostelium discoideum: An Alternative Nonanimal Model for Developmental Toxicity Testing.

Publications

Dictyostelium discoideum: An Alternative Nonanimal Model for Developmental Toxicity Testing.

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