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<figure><img src="image_3.png" /><figcaption>Figures</figcaption></figure>
Figure 1

**L. stagnalis NR relationships.** Phylogenetic relationships among L. stagnalis NR families: ML tree (Jones-Taylor-Thornton substitution model, Gamma distributed substitution rate with 4 categories, statistical support from 1,000 bootstraps) based on ClustalW-aligned concatenated sequences of the DBD and most conserved part of the LBD of L. stagnalis NRs and their BLAST best-hits (alignment available upon request). Family NR0 was excluded from this analysis as it could not be aligned with DBD-only and LBD-only sequences. The triangles represent collapsed clades, with the number of branches/sequences indicated in brackets (see details on the full tree provided in Supplementary Figure SF2).
<figure><img src="image_4.png" /><figcaption>Figure 2</figcaption></figure>

**Phylogenetic analysis of 2DBD-NRs relationships across metazoans.** Maximum Likelihood (ML) tree (same parameters as in Figure 1; values below branches indicate statistical support based on 1,000 bootstraps) based on the ClustalW-aligned concatenated conserved domains (DBD1, DBD2, and LBD) of the two sequences found in the genome of *L. stagnalis* (present study, indicated by red dots), and 60 sequences identified by Wu and Loverde (2023) from various lineages (Brachiopoda, Rotifera,
Platyhelminthes, Annelida, Mollusca, Nematoda, Echinodermata, Cephalochordata, indicated by the different colours) and having the three domains complete (as verified using Conserved Domains DataBase). The different NR7 groups referred to in the text are indicated on the right in blue, red and grey. Alignment available upon request. See also Supplementary Figure SF3 for detailed clade composition).

<figure><img src="image_5.png" /><figcaption>**Figure 3**</figcaption></figure>

**Gastropod proteome orthology.** Left: phylogenetic relationships among 14 gastropod species (topology taken from ¹³⁹) used for gene orthology assessment (colour code: blue, simultaneous hermaphrodite; purple, sequential hermaphrodite: pink, separate sexed). Right: pairwise correlation of species contribution in terms of number of proteins to 29,428 orthogroups produced by OrthoFinder. Correlations are particularly strong within the set of euthyneuran species used in the analysis. The heat map gradient on the right indicates the level of (positive) correlation.
<figure><img src="image_6.png" /><figcaption>Figure 4</figcaption></figure>

Representative whole mount *in situ* hybridisation (WMISH) expression profiles for a selection of 2DBD-NRs, NRs, GPCRs and ovipostatin differentially expressed between early veliger stage and adult reproductive organs (paired albumen and prostate glands). The expression patterns of these receptors and ovipostatin in early-veliger-stage larvae are broad, and generally similar; all genes are robustly expressed in ectodermal cells covering the foot (black arrowheads), surrounding the stomodaeum (white
asterisk), the mantle (white arrowhead) and the cephalic lobes (yellow arrowheads). In adult tissues ovipostatin is highly expressed in the prostate and effectively undetectable in the albumen gland as expected. Qualitative differences are also evident for many of the receptors surveyed, and hint at possible roles in maintaining these simultaneously active reproductive organs. Representative scale bars are 100 mm.

<figure><img src="image_7.png" /><figcaption>Figure 5</figcaption></figure>

**WMISH patterns of 2 2DBD-NRs during early development.** Both of these receptors display dynamic, asymmetric patterns of expression prior to and during gastrulation. Expression of GSLYST00003556001 appears to largely be restricted to nuclei, other than 2 large prominent cells in late blastulae. Conversely, transcripts of GSLYST00001311001 are distributed throughout the cytoplasm. Both genes are restricted to the vegetal hemisphere in early gastrulae and are concentrated around the blastopore.

**Supplementary Files**

This is a list of supplementary files associated with this preprint. Click to download.

* SuplTFSCIREP.pdf