Annelid adult cell type diversity and their pluripotent cellular origins

Abstract

Annelids are a broadly distributed, highly diverse, economically and environmentally important group of animals. Most species can regenerate missing body parts, and many are able to reproduce asexually. Therefore, many annelids can generate all adult cell types in adult stages. However, the putative adult stem cell populations involved in these processes, as well as the diversity of adult cell types generated by them, are still unknown. Here, we recover 75,218 single cell transcriptomes of Pristina leidyi, a highly regenerative and asexually-reproducing freshwater annelid. We characterise all major annelid adult cell types, and validate many of our observations by HCR in situ hybridisation. Our results uncover complex patterns of regionally expressed genes in the annelid gut, as well as neuronal, muscle and epidermal specific genes. We also characterise annelid-specific cell types such as the chaetal sacs and globin+ cells, and novel cell types of enigmatic affinity, including a vigilin+ cell type, a lumbrokinase+ cell type, and a diverse set of metabolic cells. Moreover, we characterise transcription factors and gene networks that are expressed specifically in these populations. Finally, we uncover a broadly abundant cluster of putative stem cells with a pluripotent signature. This population expresses well-known stem cell markers such as vasa, piwi and nanos homologues, but also shows heterogeneous expression of differentiated cell markers and their transcription factors. In these piwi+ cells, we also find conserved expression of pluripotency regulators, including multiple chromatin remodelling and epigenetic factors. Finally, lineage reconstruction analyses reveal the existence of differentiation trajectories from piwi+ cells to diverse adult types. Our data reveal the cell type diversity of adult annelids for the first time and serve as a resource for studying annelid cell types and their evolution. On the other hand, our characterisation of a piwi+ cell population with a pluripotent stem cell signature will serve as a platform for the study of annelid stem cells and their role in regeneration.

Insights into deuterostome evolution from the biphasic transcriptional programme of hemichordates

“…Using the whole temporal transcriptome datasets of each species, we subsetted these datasets to retrieve only common one-to-one orthologs and calculated distance and correlation metrics (Pearson correlation, Spearman correlation, and Jensen-Shannon Divergence) between pairs of transcriptomic stages. We calculated the average Jensen-Shannon Divergence using bootstrapping of 1000 iterations with replacement, and also performed co-occurrence analysis (Levy et al 2021; Álvarez-Campos et al 2023) using all genes and bootstrapping with replacement using the concatenate of the two datasets.…”

Allometry of cell types in planarians by single cell transcriptomics

“…Briefly, the longest isoforms per gene were extracted using gffread on an AGAT-standardised version of the GFF3 annotation file and the genome sequence FASTA. These sequences were translated into proteins using TransDecoder (https://github.com/TransDecoder/TransDecoder) using evidence from BLAST reciprocal best hits against the UniProt database and hmmer queried against the PFAM database, as previously described ( 37 ). Protein sequences were further annotated using (i) eggNOG against the protein set of all metazoa ( 59 ), (ii) InterProScan against the PFAM, SFAM, and Panther databases, (iii) BLAST reciprocal best hits against the UniProt database, and (iv) assigning co-orthologues from a set of model organism species using OrthoFinder ( 60 ).…”

“…We then processed these reads through our SPLiT-seq analysis pipeline ( 21, 37 ). The final dataset contained 28,738 cells, consistent with the ∼19Kx2 cells observed in each of the 2 FACS sorted samples.…”

Annelids as models of germ cell and gonad regeneration

“…Cells in the growth zone express germline/multipotency markers (Gazave et al, 2013; Özpolat & Bely, 2016). Because of the molecular signature similarity of the growth zone to germ cells, and the pluripotent nature of these stem cells as a population (Álvarez‐Campos et al, 2023a), they cannot be ruled out as a potential secondary source of germ cells in addition to the primordial germ cells established during embryogenesis (Özpolat et al, 2017; Rebscher, 2014; Rebscher et al, 2012). As the new segments are being established, new gonads or germ cells could be made as a part of the posterior growth process.…”

“…In regeneration, most new cells and tissues are thought to arise from cells nearby the wound site (Bely, 2014; Bely et al, 2014; Bideau et al, 2021, 2023; Boilly, 1967, 1969; Kostyuchenko & Kozin, 2021; Planques et al, 2019; Ribeiro et al, 2021). New cells in the zones of growth (especially posterior growth) are thought to arise from a population of cells that altogether have pluripotency but subpopulations have more limited potencies, and the individual potencies of these cells are yet to be determined (Álvarez‐Campos et al, 2023a; Balavoine, 2014; Gazave et al, 2013).…”