ABSTRACT Single-cell RNA transcriptomic platforms have significantly contributed to our understanding of tissue heterogeneity as well as of developmental and cellular differentiation trajectories. They also provide an opportunity to map an organism’s response to environmental cues with high resolution and unbiasedly identify the cell types that are the most transcriptionally sensitive to exposures. Here, we applied single nucleus RNA-seq experimental and computational approaches to C. elegans to establish the transcriptome of the adult nematode and comprehensively characterize the transcriptional impact of ethanol as a model environmental exposure on the entire organism at cell type-resolution over several generations. Clustering, tissue and phenotype enrichment, and gene ontology analyses identified 31 clusters representing a diverse number of adult cell types, including those from syncytial and multi-nucleated tissues which are difficult to assess by single cell RNA-seq, such as the mitotic and meiotic germline, hypodermal cells, and the intestine. We applied this method to identify the impact of inter- and trans-generational exposure to two human-relevant doses of alcohol. Cell type proportions were not significantly altered by ethanol. However, Euclidean distance analysis identified several germline, striated muscle, and neuronal clusters as being major transcriptional targets of ethanol at both the F1 and F3 generations although the relative order of clusters changed between generations. The impact on germline clusters was further confirmed by phenotypic enrichment analysis as well as functional validation, namely a remarkable inter- and trans-generational increase in germline apoptosis, aneuploidy, and embryonic lethality. Together, snRNA-seq of the adult C. elegans represents a powerful approach for the detailed examination of an adult organism’s response to environmental cues.

Single-nucleus resolution mapping of the adult C. elegans and its application to elucidate inter- and trans-generational response to alcohol