Single cell RNA sequencing of theStrongylocentrotus purpuratuslarva reveals the blueprint of major cell types and nervous system of a non-chordate deuterostome

Abstract

Identifying the molecular fingerprint of organismal cell types is key for understanding their function and evolution. Here, we use single cell RNA sequencing (scRNA-seq) to survey the cell types of the sea urchin early pluteus larva, representing an important developmental transition from non-feeding to feeding larva. We identified 21 distinct cell clusters, representing cells of the digestive, skeletal, immune, and nervous systems. Further subclustering of these revealed a highly detailed portrait of cell diversity across the larva, including the identification of 12 distinct neuronal cell types. Moreover, we corroborated co-expression of key regulatory genes previously shown to drive sea urchin gene regulatory networks, and revealed additional domains in which these regulatory networks are likely to function within the larva. Lastly, we recovered a neuronal cell type co-expressing Pdx-1 and Brn1/2/4, which had previously been shown to share similar gene expression with vertebrate pancreas. Our results extend this finding, revealing twenty transcription factors shared by this population of neurons in sea urchin and vertebrate pancreatic cells. Using differential expression results from Pdx-1 knockdown experiments, we generate a draft of the Pdx-1 regulatory network in these cells, and hypothesize this network was present in an ancestral deuterostome neuron before being co-opted into the pancreas developmental lineage in vertebrates.

Integrating single cell transcriptomics and volume electron microscopy confirms the presence of pancreatic acinar-like cells in sea urchins

“…It is important to stress out that previous studies from our group demonstrated that sea urchin could potentially represent an intermediate stage in pancreas evolution, since the larva contains, apart from the endodermally-derived endocrine and exocrine-like cell types (Perillo et al, 2016). Moreover we recently provided evidence that sea urchin larvae possess an additional neuronal population with a pancreatic-like gene toolkit, whose differentiation is Pdx1dependent, and was probably co-opted by modern endocrine pancreatic cells (Paganos et al, 2021). This scenario is also supported by the molecular and functional similarities of mammalian neurons and β pancreatic cells (Arntfield and van der Kooy, 2011).…”

“…The Single cell RNA data analyzed in this study originate from the study by Paganos et al (2021) as they have been deposited in Dryad under the unique identifier https://doi.org/ 10.5061/dryad.n5tb2rbvz. Subclustering analysis has been performed as described in Paganos et al (2021) using the Seurat scRNA-seq R package (Butler et al, 2018;Stuart et al, 2019).…”

“…The Single cell RNA data analyzed in this study originate from the study by Paganos et al (2021) as they have been deposited in Dryad under the unique identifier https://doi.org/ 10.5061/dryad.n5tb2rbvz. Subclustering analysis has been performed as described in Paganos et al (2021) using the Seurat scRNA-seq R package (Butler et al, 2018;Stuart et al, 2019). Subclustering analysis was performed by selecting and subsetting the cell type families of endodermal origin (esophagus, cardiac sphincter, exocrine pancreas-like, stomach 1, 2, and 3, pyloric sphincter, intestine and anus).…”

“…Single cell RNA sequencing has been shown to be a great resource in analyzing fine developmental events as well as identifying complex molecular signatures during sea urchin embryogenesis (Massri et al, 2021;Paganos et al, 2021). Previously, we have used single cell RNA sequencing originating from 3 dpf S. purpuratus larvae to reconstruct the cell type family composition of the S. purpuratus pluteus larva (Paganos et al, 2021).…”

“…Nowadays, technological advances in transcriptomics and imaging have increased the ease with which the combination of molecular fingerprint plus high-resolution morphology of cell types enable the tackling of such complex questions. For instance, single cell RNA sequencing (scRNA-seq) applied on organisms including both vertebrates and invertebrates, such as sponges, planarians, mollusks, cnidarians and echinoderms has led to the recognition of novel cell type families and the understanding of developmental and gene regulatory processes in these animals at an unprecedented level of detail (Fincher et al, 2018;Sebe-Pedros et al, 2018;Cao et al, 2019;Tabula Muris, 2020;Chari et al, 2021;Musser et al, 2021;Paganos et al, 2021;Salamanca-Díaz et al, 2022). Furthermore, single cell inventories have also allowed the reevaluation of developmental and evolutionary relationships of animals' cell types.…”