Journal: bioRxiv

Article Title: Cellular and transcriptional trajectories of neural fate specification in sea anemone uncover two modes of adult neurogenesis

doi: 10.1101/2025.06.06.655355

Figure Lengend Snippet: A Active adult neurogenesis highlighted by Elav>KikGR reporter photoconversion and in situ imaging (see Supplementary Fig. 1). At 2,4 and 7 days post-photoconversion, neural cells with the new green reporter signal and lacking the day 0 photoconverted reporter signal are newly-expressing-only cells (green cells, represented in blue throughout the figures). Cells with the photoconverted tracer either also display new KikGR expression (ongoing expression, orange cells) or lack new KikGR expression (progeny cells, red cells). B Elav>KikGR + body wall nerve net at 4 days post-photoconversion with 5X global field of view ( left ) and 40X single-cell zoom-in ( right ) images. Photoconverted KikGR day 0 tracer signal is shown in red, and new KikGR expression in cyan. Insets highlight green neurons (white arrows), orange neurons (orange arrows) and red neurons (magenta arrows). Scale bars are 500 µm (left), 50 µm (right) and 10 µm (insets). C Proportion of green, orange and red Elav>KikGR + cells detected across raw images in the oral and aboral regions (with hachure pattern) at 4 days post-photoconversion. For each image, cell counts are normalized by the total number of Elav>KikGR + cells detected (n=3 animals, mean ±S.D.). D Adult generation of Elav + neurons from Elav -negative cells across the body wall nerve net. Data is provided as Source Data File 1.

Article Snippet: For the FoxL2>KikGR , SoxC>KikGR and Elav>KikGR reporter line generation, plasmid constructs generated from a pNvT-MHC::mCh vector backbone (a gift from Ulrich Technau, Addgene plasmid #67943 ) with the KikGR fluorophore (from the pCAG::KikGR vector, a gift from Anna-Katerina Hadjantonakis, Addgene plasmid #32608 ).

Techniques: In Situ, Imaging, Expressing

Journal: bioRxiv

Article Title: Cellular and transcriptional trajectories of neural fate specification in sea anemone uncover two modes of adult neurogenesis

doi: 10.1101/2025.06.06.655355

Figure Lengend Snippet: A Molecular identification of newly-generated Elav + neurons. Plate-based scRNA-Seq (MARS-Seq protocol) allows recording of reporter status post-photoconversion (green, orange or red cell) by flow cytometry for each cell sampled from Elav>KikGR + adult body regions (body wall with mesenteries) at 2,4 and 7 days post-photoconversion. scRNA-Seq transcriptional profiles are then matched to each cell’s reporter status color (Supplementary Fig. 3 and 4). B UMAP projection of the 1335 peptidergic neurons from the 3006 Elav>KikGR + sampled cells from 2, 4 and 7 days post-photoconversion with their cluster identifier. C Scaled neural marker gene expression across Elav>KikGR + peptidergic neuron clusters. Broadly expressed neural markers and neuropeptide processing genes ( top ) and specifically expressed neuropeptide precursor genes from , ( bottom ). Dot size represents the percentage of cells in this cluster which express the gene. Neural cluster identifiers as in B . See Supplementary Fig. 5 for expression across all Elav>KikGR + cells. D Elav + peptidergic neuron scRNA-seq UMAP projection highlighting the overlapping identity distribution between the previously-labeled peptidergic neurons (orange and red cells) ( left ), and the newly-expressing neurons (green) ( right ) at 7 days post-photoconversion. E Cell counts by reporter status (green vs. red and orange cells) across peptidergic cell clusters in 7 days photoconverted Elav>KikGR + animals showing a Pearson’s correlation R of 0.95. Cluster identifiers as in B-C . Data is provided as Source Data File 2.

Article Snippet: For the FoxL2>KikGR , SoxC>KikGR and Elav>KikGR reporter line generation, plasmid constructs generated from a pNvT-MHC::mCh vector backbone (a gift from Ulrich Technau, Addgene plasmid #67943 ) with the KikGR fluorophore (from the pCAG::KikGR vector, a gift from Anna-Katerina Hadjantonakis, Addgene plasmid #32608 ).

Techniques: Generated, Flow Cytometry, Marker, Gene Expression, Expressing, Labeling

Journal: bioRxiv

Article Title: Cellular and transcriptional trajectories of neural fate specification in sea anemone uncover two modes of adult neurogenesis

doi: 10.1101/2025.06.06.655355

Figure Lengend Snippet: SoxC>KikGR + (A) and FoxL2>KikGR+ (C) adult polyp body wall at 7 days post reporter photoconversion. Global field of view (5X, left) and high magnification image (40X extended depth of focus ( A ) or single image ( C ), right) with photoconverted tracer in red and new signal in cyan. SoxC driver insets highlight cnidocytes (asterisks) and neurons (arrows) with either only the photoconverted signal (in magenta), or only the new expression (in white), as well as secretory cells with both photoconverted and new signal (orange arrowhead). FoxL2 driver insets highlight neurons with both photoconverted and new signal (orange arrows) and neurons with only photoconverted KikGR signal (magenta arrows). Scale bars are 500 µm ( left ), 50 µm ( right ), and 10 µm in the insets. B,D Proportion of red, orange and green cell bodies detected in the body wall at 7 days post-photoconversion with the SoxC ( B ) and FoxL2 ( D ) drivers. Aboral end body wall images are indicated with a hachure pattern (n=5-6 for the SoxC driver and n=2-3 for the FoxL2 driver, average proportion +/- S.D.). E scRNA-Seq UMAP projection of 12,352 KikGR + cells from the Elav , FoxL2 and SoxC drivers at 2, 4 and 7 days post-photoconversion, colored by broad cluster identity. F Marker gene expression across clusters. For each gene, expression is normalized to the highest expression across clusters. Colors correspond to broad cluster annotations, as in E . Dot size represents the proportion of cells expressing the gene in the cluster. Marker genes names are curated from , , , ,41, , see Supplementary File 1. G Cell identity distribution by driver and FACS gating strategy across the SoxC , FoxL2 and Elav drivers. For the FoxL2 driver, the KikGR+ no epi. FACS gate depleted the epithelial epidermis cell fraction. Data is provided as Source Data File 3.

Article Snippet: For the FoxL2>KikGR , SoxC>KikGR and Elav>KikGR reporter line generation, plasmid constructs generated from a pNvT-MHC::mCh vector backbone (a gift from Ulrich Technau, Addgene plasmid #67943 ) with the KikGR fluorophore (from the pCAG::KikGR vector, a gift from Anna-Katerina Hadjantonakis, Addgene plasmid #32608 ).

Techniques: Expressing, Marker, Gene Expression

Journal: bioRxiv

Article Title: Cellular and transcriptional trajectories of neural fate specification in sea anemone uncover two modes of adult neurogenesis

doi: 10.1101/2025.06.06.655355

Figure Lengend Snippet: A Putative SoxC +/ FoxL2 + Piwi1 + progenitors differentiating into Elav + peptidergic neurons highlights predicted differential neuron reporter status between the FoxL2 and SoxC drivers, with only red and orange neurons expected from the FoxL2 driver while red, orange and green neurons would be recovered from the SoxC driver at 2, 4 and 7 days post reporter photoconversion. Green neurons are shown in blue. B Green, orange and red cell proportions for Elav + peptidergic neurons recovered from the FoxL2 (left) and SoxC (right) drivers at 2, 4 and 7 days post-photoconversion. Elav + peptidergic neurons from the FoxL2 driver are red or orange. Green, orange and red SoxC driver neurons are recovered, with the arrow indicating the increased fraction of newly generated progeny neurons (green and orange cells) at later timepoints. C Schematics for two scenarios of peptidergic neuron development: distinct Elav + neuron clusters represent (top) temporal differentiation intermediates or (bottom) terminally differentiated populations concomitantly generated. As the SoxC driver transiently labels differentiating cells, tracking the ratio between newly arising green+orange neurons and previously-generated red neurons recovered from each peptidergic cluster can discriminate between the two scenarios. This ratio is similar across clusters that are terminal states (bottom) but not for clusters that are transient intermediates (top). Term.: terminally differentiated neural state. Letters indicate neuron clusters. D SoxC>KikGR + peptidergic neuron clusters distribution at 7 days post-photoconversion highlights a similar fraction of green and orange cells relative to the red cells across all peptidergic clusters. E Elav + peptidergic cluster cell counts recovered from the SoxC versus the FoxL2 driver ( KikGR+ cells, all days post-photoconversion), in blue (R =0.65), with digestive secretory cells in orange, and other neural and secretory cell clusters for comparison. F Elav + peptidergic neuron development model, with differentiation into distinct peptidergic neural subtypes from peptidergic-committed Piwi1 + multipotent cells within 4 days in adults. Data is provided as Source Data File 5.

Article Snippet: For the FoxL2>KikGR , SoxC>KikGR and Elav>KikGR reporter line generation, plasmid constructs generated from a pNvT-MHC::mCh vector backbone (a gift from Ulrich Technau, Addgene plasmid #67943 ) with the KikGR fluorophore (from the pCAG::KikGR vector, a gift from Anna-Katerina Hadjantonakis, Addgene plasmid #32608 ).

Techniques: Generated, Comparison