For more than 100 years, the fruit fly Drosophila melanogaster has been one of the most studied model organisms. Here, we present a single-cell atlas of the adult fly, Tabula Drosophilae, that includes 580,000 nuclei from 15 individually dissected sexed tissues as well as the entire head and body, annotated to >250 distinct cell types. We provide an in-depth analysis of cell type–related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types between tissues, such as blood and muscle cells, reveals rare cell types and tissue-specific subtypes. This atlas provides a valuable resource for the Drosophila community and serves as a reference to study genetic perturbations and disease models at single-cell resolution. Description Cell type diversity in a whole fly The fruit fly Drosophila melanogaster has served as a premier model organism for discovering fundamental and evolutionarily conserved biological mechanisms. Combining recent advances in single-cell sequencing with powerful fly genetic tools holds great promise for making further discoveries. Li et al. present a single-cell atlas of the entire adult fly that includes 580,000 cells and more than 250 annotated cell types. Cells from the head and body recapitulated cell types from 15 dissected tissues. In-depth analyses revealed rare cell types, cell-type-specific gene signatures, and sexual dimorphism. This atlas provides a resource for the Drosophila community to study genetic perturbations and diseases at single-cell resolution. —BAP A single-nucleus transcriptomic map reveals more than 250 distinct cell types in the entire adult Drosophila melanogaster. INTRODUCTION Drosophila melanogaster has had a fruitful history in biological research because it has contributed to many key discoveries in genetics, development, and neurobiology. The fruit fly genome contains ~14,000 protein-coding genes, ~63% of which have human orthologs. Single-cell RNA-sequencing has recently been applied to multiple Drosophila tissues and developmental stages. However, these data have been generated by different laboratories on different genetic backgrounds with different dissociation protocols and sequencing platforms, which has hindered the systematic comparison of gene expression across cells and tissues. RATIONALE We aimed to establish a cell atlas for the entire adult Drosophila with the same genetic background, dissociation protocol, and sequencing platform to (i) obtain a comprehensive categorization of cell types, (ii) integrate single-cell transcriptome data with existing knowledge about gene expression and cell types, (iii) systematically compare gene expression across the entire organism and between males and females, and (iv) identify cell type–specific markers across the entire organism. We chose single-nucleus RNA-sequencing (snRNA-seq) to circumvent the difficulties of dissociating cells that are embedded in the cuticle (e.g., sensory neurons) or that are multinucleated (e.g., muscle cells). We took two complementary strategies: sequencing nuclei from dissected tissues to know the identity of the tissue source and sequencing nuclei from the entire head and body to ensure that all cells are sampled. Experts from 40 laboratories participated in crowd annotation to assign transcriptomic cell types with the best knowledge available. RESULTS We sequenced 570,000 cells using droplet-based 10x Genomics from 15 dissected tissues as well as whole heads and bodies, separately in females and males. We also sequenced 10,000 cells from dissected tissues using the plate-based Smart-seq2 platform, providing deeper coverage per cell. We developed reproducible analysis pipelines using NextFlow and implemented a distributed cell-type annotation system with controlled vocabularies in SCope. Crowd-based annotations of transcriptomes from dissected tissues identified 17 main cell categories and 251 detailed cell types linked to FlyBase ontologies. Many of these cell types are characterized for the first time, either because they emerged only after increasing cell coverage or because they reside in tissues that had not been previously subjected to scRNA-seq. The excellent correspondence of transcriptomic clusters from whole body and dissected tissues allowed us to transfer annotations and identify a few cuticular cell types not detected in individual tissues. Cross-tissue analysis revealed location-specific subdivisions of muscle cells and heterogeneity within blood cells. We then determined cell type–specific marker genes and transcription factors with different specificity levels, enabling the construction of gene regulatory networks. Finally, we explored sexual dimorphism, finding a link between sex-biased expression and the presence of doublesex, and investigated tissue dynamics through trajectory analyses. CONCLUSION Our Fly Cell Atlas (FCA) constitutes a valuable resource for the Drosophila community as a reference for studies of gene function at single-cell resolution. All the FCA data are freely available for further analysis through multiple portals and can be downloaded for custom analyses using other single-cell tools. The ability to annotate cell types by sequencing the entire head and body will facilitate the use of Drosophila in the study of biological processes and in modeling human diseases at a whole-organism level with cell-type resolution. All data with annotations can be accessed from www.flycellatlas.org, which provides links to SCope, ASAP, and cellxgene portals. Tabula Drosophilae. In this single-cell atlas of the adult fruit fly, 580,000 cells were sequenced and >250 cell types were annotated. They are from 15 individually dissected sexed tissues as well as the entire head and body. All data are freely available for visualization and download, with featured analyses shown at the bottom right.

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Fly Cell Atlas: A single-nucleus transcriptomic atlas of the adult fruit fly

Calcium carbonate (CaCO3) biomineralizing organisms have played major roles in the history of life and the global carbon cycle during the past 541 Ma. Both marine diversification and mass extinctions reflect physiological responses to environmental changes through time. An integrated understanding of carbonate biomineralization is necessary to illuminate this evolutionary record and to understand how modern organisms will respond to 21st century global change. Biomineralization evolved independently but convergently across phyla, suggesting a unity of mechanism that transcends biological differences. In this review, we combine CaCO3 skeleton formation mechanisms with constraints from evolutionary history, omics, and a meta-analysis of isotopic data to develop a plausible model for CaCO3 biomineralization applicable to all phyla. The model provides a framework for understanding the environmental sensitivity of marine calcifiers, past mass extinctions, and resilience in 21st century acidifying oceans. Thus, it frames questions about the past, present, and future of CaCO3 biomineralizing organisms.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8906573

Biomineralization: Integrating mechanism and evolutionary history

The ability to obtain single cell transcriptomes for stable cell types and dynamic cell states is ushering in a new era for biology. We created the Tabula Drosophilae , a single cell atlas of the adult fruit fly which includes 580k cells from 15 individually dissected sexed tissues as well as the entire head and body. Over 100 researchers from the fly community contributed annotations to >250 distinct cell types across all tissues. We provide an in-depth analysis of cell type-related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types that are shared between tissues, such as blood and muscle cells, allowed the discovery of rare cell types and tissue-specific subtypes. This atlas provides a valuable resource for the entire Drosophila community and serves as a comprehensive reference to study genetic perturbations and disease models at single cell resolution.

/pdf/fly-cell-atlas-a-single-cell-transcriptomic-atlas-of-the-vvv9i3x3vz.pdf

Fly Cell Atlas: a single-cell transcriptomic atlas of the adult fruit fly

Evolutionary cell type mapping with single-cell genomics.

Abstract Venoms have evolved >100 times in all major animal groups, and their components, known as toxins, have been fine-tuned over millions of years into highly effective biochemical weapons. There are many outstanding questions on the evolution of toxin arsenals, such as how venom genes originate, how venom contributes to the fitness of venomous species, and which modifications at the genomic, transcriptomic, and protein level drive their evolution. These questions have received particularly little attention outside of snakes, cone snails, spiders, and scorpions. Venom compounds have further become a source of inspiration for translational research using their diverse bioactivities for various applications. We highlight here recent advances and new strategies in modern venomics and discuss how recent technological innovations and multi-omic methods dramatically improve research on venomous animals. The study of genomes and their modifications through CRISPR and knockdown technologies will increase our understanding of how toxins evolve and which functions they have in the different ontogenetic stages during the development of venomous animals. Mass spectrometry imaging combined with spatial transcriptomics, in situ hybridization techniques, and modern computer tomography gives us further insights into the spatial distribution of toxins in the venom system and the function of the venom apparatus. All these evolutionary and biological insights contribute to more efficiently identify venom compounds, which can then be synthesized or produced in adapted expression systems to test their bioactivity. Finally, we critically discuss recent agrochemical, pharmaceutical, therapeutic, and diagnostic (so-called translational) aspects of venoms from which humans benefit.

Modern venomics—Current insights, novel methods, and future perspectives in biological and applied animal venom research

A stony coral cell atlas illuminates the molecular and cellular basis of coral symbiosis, calcification, and immunity.

The metabotropic gamma-aminobutyric acid B receptor (GABABR) is a G protein-coupled receptor that mediates neuronal inhibition by the neurotransmitter GABA. While GABABR-mediated signalling has been suggested to play central roles in neuronal differentiation and proliferation across evolution, it has mostly been studied in the mammalian brain. Here, we demonstrate that ectopic activation of GABABR signalling affects neurogenic functions in the sea anemone Nematostella vectensis. We identified four putative Nematostella GABABR homologues presenting conserved three-dimensional extracellular domains and residues needed for binding GABA and the GABABR agonist baclofen. Moreover, sustained activation of GABABR signalling reversibly arrests the critical metamorphosis transition from planktonic larva to sessile polyp life stage. To understand the processes that underlie the developmental arrest, we combined transcriptomic and spatial analyses of control and baclofen-treated larvae. Our findings reveal that the cnidarian neurogenic programme is arrested following the addition of baclofen to developing larvae. Specifically, neuron development and neurite extension were inhibited, resulting in an underdeveloped and less organized nervous system and downregulation of proneural factors including NvSoxB(2), NvNeuroD1 and NvElav1. Our results thus point to an evolutionarily conserved function of GABABR in neurogenesis regulation and shed light on early cnidarian development. The role of the metabotropic gamma-aminobutyric acid B receptor (GABABR) in neural differentiation is well known in vertebrates. Here, the authors show that GABABRs plays a role in neurogenesis and development in the sea anemone Nematostella vectensis.

Ectopic activation of GABAB receptors inhibits neurogenesis and metamorphosis in the cnidarian Nematostella vectensis.

Microbial communities confer multiple beneficial effects to their multicellular hosts. To evaluate the evolutionary and ecological implications of the animal-microbe interactions, it is essential to understand how bacterial colonization is secured and maintained during the transition from one generation to the next. However, the mechanisms of symbiont transmission are poorly studied for many species, especially in marine environments, where the surrounding water constitutes an additional source of microbes. Nematostella vectensis, an estuarine cnidarian, has recently emerged as model organism for studies on host-microbes interactions. Here, we use this model organism to study the transmission of bacterial colonizers, evaluating the contribution of parental and environmental transmission to the establishment of bacterial communities of the offspring. We induced spawning in adult male and female polyps of N. vectensis and used their gametes for five individual fertilization experiments. While embryos developed into primary polyps, we sampled each developmental stage and its corresponding medium samples. By analyzing the microbial community compositions of all samples through 16S rRNA gene amplicon sequencing, we showed that all host tissues harbor microbiota significantly different from the surrounding medium. Interestingly, oocytes and sperms are associated with distinct bacterial communities, indicating the specific vertical transmission of bacterial colonizers by the gametes. These differences were consistent among all the five families analyzed. By overlapping the identified bacterial ASVs associated with gametes, offspring and parents, we identified specific bacterial ASVs that are well supported candidates for vertical transmission via mothers and fathers. This is the first study investigating bacteria transmission in N. vectensis, and among few on marine spawners that do not brood larvae. Our results shed light on the consistent yet distinct maternal and paternal transfer of bacterial symbionts along the different life stages and generations of an aquatic invertebrate.

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Contribution of Maternal and Paternal Transmission to Bacterial Colonization in Nematostella vectensis.

Stony coral exoskeletons build the foundation to the most biologically diverse yet fragile marine ecosystems on earth, coral reefs. Understanding corals biomineralization mechanisms is therefore crucial for coral reef management and for using coral skeletons in geochemical studies. In this study, we combine in vivo and cryo-electron microscopy with single cell RNA-seq data to gain novel insights into the calcifying micro environment that facilitates biomineralization in primary polyps of the stony coral Stylophora pistillata. We show an intimate involvement of seawater in this micro environment. We further document increased tissue permeability and a highly dispersed cell packing in the tissue secreting the coral skeleton (i.e. calicoblastic). We also observe an extensive filopodial network containing carbon-rich vesicles extruding from some of the calicoblastic cells. Single cell RNA-Seq data interrogation shows that calicoblastic cells express genes involved in filopodia and vesicle structure and function. These observations provide a new conceptual framework for resolving stony corals biomineralization processes.

https://biorxiv.org/content/10.1101/2021.06.09.447817v1.full.pdf

Shani Levy

Papers

A stony coral cell atlas illuminates the molecular and cellular basis of coral symbiosis, calcification, and immunity.

Ectopic activation of GABAB receptors inhibits neurogenesis and metamorphosis in the cnidarian Nematostella vectensis.

Contribution of Maternal and Paternal Transmission to Bacterial Colonization in Nematostella vectensis.

The calcifying interface in a stony coral's primary polyp: An interplay between seawater and an extracellular calcifying space

The calcifying interface in a stony coral primary polyp: An interplay between seawater and an extracellular calcifying space.