Mating in the Closest Living Relatives of Animals Is Induced by a Bacterial Chondroitinase.

We serendipitously discovered that the marine bacterium Vibrio fischeri induces sexual reproduction in one of the closest living relatives of animals, the choanoflagellate Salpingoeca rosetta. Although bacteria influence everything from nutrition and metabolism to cell biology and development in eukaryotes, bacterial regulation of eukaryotic mating was unexpected. Here, we show that a single V. fischeri protein, the previously uncharacterized EroS, fully recapitulates the aphrodisiac-like activity of live V. fischeri. EroS is a chondroitin lyase; although its substrate, chondroitin sulfate, was previously thought to be an animal synapomorphy, we demonstrate that S. rosetta produces chondroitin sulfate and thus extend the ancestry of this important glycosaminoglycan to the premetazoan era. Finally, we show that V. fischeri, purified EroS, and other bacterial chondroitin lyases induce S. rosetta mating at environmentally relevant concentrations, suggesting that bacteria likely regulate choanoflagellate mating in nature.

A wound-induced differentiation trajectory for neurons.

Animals capable of whole-body regeneration can replace any missing cell type and regenerate fully functional new organs, including new brains, de novo. The regeneration of a new brain requires the formation of diverse neural cell types and their assembly into an organized structure with correctly wired circuits. Recent work in various regenerative animals has revealed transcriptional programs required for the differentiation of distinct neural subpopulations, however, how these transcriptional programs are initiated in response to injury remains unknown. Here, we focused on the highly regenerative acoel worm, Hofstenia miamia, to study wound-induced transcriptional regulatory events that lead to the production of neurons and subsequently a functional brain. Footprinting analysis using chromatin accessibility data on a chromosome-scale genome assembly revealed that binding sites for the Nuclear Factor Y (NFY) transcription factor complex were significantly bound during regeneration, showing a dynamic increase in binding within one hour upon amputation specifically in tail fragments, which will regenerate a new brain. Strikingly, NFY targets were highly enriched for genes with neuronal function. Single-cell transcriptome analysis combined with functional studies identified soxC+ stem cells as a putative progenitor population for multiple neural subtypes. Further, we found that wound-induced soxC expression is likely under direct transcriptional control by NFY, uncovering a mechanism for the initiation of a neural differentiation pathway by early wound-induced binding of a transcriptional regulator.

Neural architecture and regeneration in the acoel Hofstenia miamia.

The origin of bilateral symmetry, a major transition in animal evolution, coincided with the evolution of organized nervous systems that show regionalization along major body axes. Studies of Xenacoelomorpha, the likely outgroup lineage to all other animals with bilateral symmetry, can inform the evolutionary history of animal nervous systems. Here, we characterized the neural anatomy of the acoel Hofstenia miamia. Our analysis of transcriptomic data uncovered orthologues of enzymes for all major neurotransmitter synthesis pathways. Expression patterns of these enzymes revealed the presence of a nerve net and an anterior condensation of neural cells. The anterior condensation was layered, containing several cell types with distinct molecular identities organized in spatially distinct territories. Using these anterior cell types and structures as landmarks, we obtained a detailed timeline for regeneration of the H. miamia nervous system, showing that the anterior condensation is restored by eight days after amputation. Our work detailing neural anatomy in H. miamia will enable mechanistic studies of neural cell type diversity and regeneration and provide insight into the evolution of these processes.

A wound-induced differentiation trajectory for neurons.

Animals capable of whole-body regeneration can replace any missing cell type and regenerate fully-functional new organs, de novo . The regeneration of a new brain requires the formation of diverse neuronal cell types and their assembly into an organized structure and correctly-wired circuits. Recent work in various regenerative animals has revealed transcriptional programs required for the differentiation of distinct neuronal subpopulations, however how these transcriptional programs are initiated upon amputation remains unknown. Here, we focused on the highly regenerative acoel worm, Hofstenia miamia , to study wound-induced transcriptional regulatory events that lead to the production of neurons. Footprinting analysis using chromatin accessibility data on an improved genome assembly revealed that binding sites for the NFY transcription factor complex were significantly bound during regeneration, showing a dynamic increase in binding within one hour upon amputation specifically in tail fragments, which will regenerate a new brain. Strikingly, NFY targets were highly enriched for genes with neuronal functional. Single-cell transcriptome analysis combined with functional studies identified sox4 + stem cells as the likely progenitor population for multiple neuronal subtypes. Further, we found that wound-induced sox4 expression is likely under direct transcriptional control by NFY, uncovering a mechanism for how early wound-induced binding of a transcriptional regulator results in the initiation of a neuronal differentiation pathway. Highlights: A new chromosome-scale assembly for Hofstenia enables comprehensive analysis of transcription factor binding during regeneration NFY motifs become dynamically bound by 1hpa in regenerating tail fragments, particularly in the loci of neural genes A sox4 + neural-specialized stem cell is identified using scRNA-seq sox4 is wound-induced and required for differentiation of multiple neural cell types NFY regulates wound-induced expression of sox4 during regeneration.

Acoel single-cell atlas reveals expression dynamics and heterogeneity of adult pluripotent stem cells.

Adult pluripotent stem cell (aPSC) populations underlie whole-body regeneration in many distantly-related animal lineages, but how the underlying cellular and molecular mechanisms compare across species is unknown. Here, we apply single-cell RNA sequencing to profile transcriptional cell states of the acoel worm Hofstenia miamia during postembryonic development and regeneration. We identify cell types shared across stages and their associated gene expression dynamics during regeneration. Functional studies confirm that the aPSCs, also known as neoblasts, are the source of differentiated cells and reveal transcription factors needed for differentiation. Subclustering of neoblasts recovers transcriptionally distinct subpopulations, the majority of which are likely specialized to differentiated lineages. One neoblast subset, showing enriched expression of the histone variant H3.3, appears to lack specialization. Altogether, the cell states identified in this study facilitate comparisons to other species and enable future studies of stem cell fate potentials.

Acoel genome reveals the regulatory landscape of whole-body regeneration.

Whole-body regeneration is accompanied by complex transcriptomic changes, yet the chromatin regulatory landscapes that mediate this dynamic response remain unexplored. To decipher the regulatory logic that orchestrates regeneration, we sequenced the genome of the acoel worm Hofstenia miamia, a highly regenerative member of the sister lineage of other bilaterians. Epigenomic profiling revealed thousands of regeneration-responsive chromatin regions and identified dynamically bound transcription factor motifs, with the early growth response (EGR) binding site as the most variably accessible during Hofstenia regeneration. Combining egr inhibition with chromatin profiling suggests that Egr functions as a pioneer factor to directly regulate early wound-induced genes. The genetic connections inferred by this approach allowed the construction of a gene regulatory network for whole-body regeneration, enabling genomics-based comparisons of regeneration across species.

Predicted glycosyltransferases promote development and prevent spurious cell clumping in the choanoflagellate S. rosetta.

In a previous study we established forward genetics in the choanoflagellate Salpingoeca rosetta and found that a C-type lectin gene is required for rosette development (Levin et al., 2014). Here we report on critical improvements to genetic screens in S. rosetta while also investigating the genetic basis for rosette defect mutants in which single cells fail to develop into orderly rosettes and instead aggregate promiscuously into amorphous clumps of cells. Two of the mutants, Jumble and Couscous, mapped to lesions in genes encoding two different predicted glycosyltransferases and displayed aberrant glycosylation patterns in the basal extracellular matrix (ECM). In animals, glycosyltransferases sculpt the polysaccharide-rich ECM, regulate integrin and cadherin activity, and, when disrupted, contribute to tumorigenesis. The finding that predicted glycosyltransferases promote proper rosette development and prevent cell aggregation in S. rosetta suggests a pre-metazoan role for glycosyltransferases in regulating development and preventing abnormal tumor-like multicellularity.