Updated single cell reference atlas for the starlet anemone Nematostella vectensis

Abstract

Background: The recent combination of genomics and single cell transcriptomics has allowed to assess a variety of non-conventional model organisms in much more depth. Single cell transcriptomes can uncover hidden cellular complexity and cell lineage relationships within organisms. The recent developmental cell atlases of the sea anemone Nematostella vectensis, a representative of the basally branching Cnidaria, has provided new insights into the development of all cell types (Steger et al Cell Rep 40(12):111370, 2022; Sebé-Pedrós et al. Cell 173(6):1520-1534.e20). However, the mapping of the single cell reads still suffers from relatively poor gene annotations and a draft genome consisting of many scaffolds.

Results: Here we present a new wildtype resource of the developmental single cell atlas, by re-mapping of sequence data first published in Steger et al. (2022) and Cole et al. (Nat Commun 14(1):1747, 2023), to the new chromosome-level genome assembly and corresponding gene models in Zimmermann et al. (Nat Commun 14, 8270 (2023). https://doi.org/10.1038/s41467-023-44080-7 ). We expand the pre-existing dataset through the incorporation of additional sequence data derived from the capture and sequencing of cell suspensions from four additional samples: 24 h gastrula, 2d planula, an inter-parietal region of the bodywall from a young unsexed animal, and another adult mesentery from a mature male animal.

Conclusion: Our analyses of the full cell-state inventory provide transcriptomic signatures for 127 distinct cell states, of which 47 correspond to neuroglandular subtypes. We also identify two distinct putatively immune-related transcriptomic profiles that segregate between the inner and outer cell layers. Furthermore, the new gene annotation Nv2 has markedly improved the mapping on the single cell transcriptome data and will therefore be of great value for the community and anyone using the dataset.

Keywords: Cnidarian development; Neuronal inventory; Transcriptomics; scRNAseq.

Fig. 1

Updated cell atlas for the starlet sea anemone Nematostella vectensis. A Dimensional reduction cell map (UMAP) illustrating clustering of cells from the developmental (left) or adult tissue (right) subsets. Color of clustering corresponds to the legend in (b). Distribution of the entire subset is shown in the inset. B Relative contribution of each library to the clusters from the developmental (left) or tissue (right) samples. C) Dotplot representation of differentially expressed gene sets from all genes (left) and restricted to transcription factors only (right). Expression profiles are split between the two subsets. Orange scale = Adult subset; Dark slate blue scale = Developmental subset

Fig. 2

Early embryonic ectoderm matures into two distinct tissues. A Sorting according to ectodermal patterning within the transcriptomic data is evidenced by comparing expression patterns known from in situ hybridization with expression in the dimensional reduction. B Schematic illustrates the locations of the ectodermal layers in the adult. UMAP dimensional reduction of the ectodermal partition colored by cell state identity highlighting the retention of both spatial and temporal organisation within the clustering. C Relative distribution of cell cluster identities across all samples in the dataset. E Dotplot representation of differentially expressed gene sets from all genes (left) and restricted to transcription factors only (right). Expression profiles are split between the two subsets. Orange scale = Adult subset; Dark slate blue scale = Developmental subset

Fig. 3

Cell of the apical organ of planula are identifiable from transcriptomic profiles. A Sample distribution of apical organ cells shown on UMAP reduction B Dotplot of differentially expressed genes (top) and transcription factors (bottom) C Cell state identities of apical organ cells (top) and known regional markers (bottom)

Fig. 4

Inner cell layer matures into gastrodermis and derived cell types. A UMAP dimensional reduction cell plot, colored by sample origin. Inset: included partitions. Dashed arrow: maturation pathway. CM: circular muscle; PM: parietal muscle; MR: mesentery retractor muscle; ImM: intermuscular membrane. B Bar plot representation of fraction of cells from each cluster (colours) within each sample (bars). C Dot plot representation of expression profiles of up-regulated genes (left) and up-regulated transcription factors (right) across each cluster. Expression separated between cells of the developmental series (dark slate blue scale) and the adult tissue series (orange scale). Grey indicates average scaled expression of 0 or below. See Supplementary material for full gene lists

Fig. 5

Sub-states of retractor muscle cells from both germ layers can be resolved. A Retractor muscle partition within the entire dataset (inset). UMAP representation of four unique cluster states. B Bar plot showing the log count (C) of cells from each sample contributing to the partition. C Dot plot of gene expression separated between partition cluster cells from the developmental (dark slate blue gradient) or adult tissue (orange gradient) samples. Expression profiles of the differentially expressed genes across the partition (Signature Genes) and each partition cluster (Top 5 All Genes), and its regulatory profile (box)

Fig. 6

Cnidocyte specification pathways are recovered. A Cell plot indicating the clusters of the cnidocyte partition (inset). B Distribution of sample contribution to each identified cluster. C Specific toxin profiles associated with cnidocyte subtypes. D Dotplot of marker expression (box) and differentially up-regulated genes of each cluster. Expression separated between cells of the developmental series (Dark slate blue scale) and the adult tissue series (orange scale). Grey indicates average scaled expression of 0 or below. See Supplementary material for full gene lists

Fig. 7

Neuroglandular precursor cells can be identified exiting from the mitotic cycle and derive from both germ layers. A Identified clusters. (inset) Partitions included in the analyses: Putative stem cells (red) and primary germ cells (blue) B Distribution of cells from each sample included in the cluster: absolute cell counts C expression profiles of mitotic markers of DNA-synthesis (PCNA) and division (NUSAP-like-1), and NPC marker soxC and pre-neural marker nanos1. D Dotplot expression of top marker genes (left) and differentially expressed transcription factors (right) from each cluster. Expression separated between cells of the developmental series (Dark slate blue scale) and the adult tissue series (orange scale). Grey indicates average scaled expression of 0 or below. See Supplementary material for full gene lists

Fig. 8

Neuroglandular derivatives include cells of multiple related cell types. A Dimensional reduction cell plot (UMAP) showing clustering of the neuroglandular partition (inset). B Bar plot of relative contribution of each cell state across all samples

Fig. 9

Neuroglandular derivatives include cells of multiple related cell types, continued. Dot plot of gene expression of the top state-specific gene sets from all genes (top) and the set of upregulated transcription factors (bottom). Expression separated between cells of the developmental series (dark slate blue scale) and the adult tissue series (orange scale). Grey indicates average scaled expression of 0 or below. See Additional file 1 for full gene lists