Anal and caudal fins in the control and treated fish at day 0 and day 5 (A) and details of fin rays in the developing sword and gonopodium (B). (A) In the control fish, no difference between anal fin (CG) and ventral caudal fin (CV) was observed at day 0 and day 5 (scale bar = 1 mm). In testosterone-treated fish, initiation of the transformation into a gonopodium (TG) and sword (TV) was apparent at day 5. (B) The gonopodium developed from rays 3, 4, and 5 in anal fin, and the sword developed from V7 to V10 (ventral). Tissues from dorsal rays (D7–D10) and middle rays (V1, V2, D1 and D2) were used for RNA sequencing. *T stands for treated, C for nontreated (control), V for ventral caudal rays (sword), M for middle caudal rays, D for dorsal caudal rays.

Anal and caudal fins in the control and treated fish at day 0 and day 5 (A) and details of fin rays in the developing sword and gonopodium (B). (A) In the control fish, no difference between anal fin (CG) and ventral caudal fin (CV) was observed at day 0 and day 5 (scale bar = 1 mm). In testosterone-treated fish, initiation of the transformation into a gonopodium (TG) and sword (TV) was apparent at day 5. (B) The gonopodium developed from rays 3, 4, and 5 in anal fin, and the sword developed from V7 to V10 (ventral). Tissues from dorsal rays (D7–D10) and middle rays (V1, V2, D1 and D2) were used for RNA sequencing. *T stands for treated, C for nontreated (control), V for ventral caudal rays (sword), M for middle caudal rays, D for dorsal caudal rays.

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Swords are exaggerated male ornaments of swordtail fishes that have been of great interest to evolutionary biologists ever since Darwin described them in the Descent of Man (1871). They are a novel sexually selected trait derived from modified ventral caudal fin rays and are only found in the genus Xiphophorus. Another phylogenetically more widespr...

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... Moreover, because the same V mem dynamics can be produced by many different ion channel combinations, and because bioelectric states propagate their influence across tissue distance during morphogenesis (Chernet and Levin, 2014;Pai et al., 2020), evolution is free to swap out channels and explore the bioelectrical state space: simple mutations in electrogenic genes can exert very long-range, highly coordinated changes in anatomy. Indeed, the KCNH8 ion channel and a connexin were identified in the transcriptomic analysis of the evolutionary shift between two functionally different morphologies of fin structures in fish (Kang et al., 2015). The evolutionary significance of bioelectric controls can also be seen across lineages, as some viruses evolved to carry ion channel and gap junction (Vinnexin) genes that enable them to hijack bioelectric machinery used by their target cells (Shimbo et al., 1996;Hover et al., 2017). ...
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Synthetic biology and bioengineering provide the opportunity to create novel embodied cognitive systems (otherwise known as minds) in a very wide variety of chimeric architectures combining evolved and designed material and software. These advances are disrupting familiar concepts in the philosophy of mind, and require new ways of thinking about and comparing truly diverse intelligences, whose composition and origin are not like any of the available natural model species. In this Perspective, I introduce TAME—Technological Approach to Mind Everywhere—a framework for understanding and manipulating cognition in unconventional substrates. TAME formalizes a non-binary (continuous), empirically-based approach to strongly embodied agency. TAME provides a natural way to think about animal sentience as an instance of collective intelligence of cell groups, arising from dynamics that manifest in similar ways in numerous other substrates. When applied to regenerating/developmental systems, TAME suggests a perspective on morphogenesis as an example of basal cognition. The deep symmetry between problem-solving in anatomical, physiological, transcriptional, and 3D (traditional behavioral) spaces drives specific hypotheses by which cognitive capacities can increase during evolution. An important medium exploited by evolution for joining active subunits into greater agents is developmental bioelectricity, implemented by pre-neural use of ion channels and gap junctions to scale up cell-level feedback loops into anatomical homeostasis. This architecture of multi-scale competency of biological systems has important implications for plasticity of bodies and minds, greatly potentiating evolvability. Considering classical and recent data from the perspectives of computational science, evolutionary biology, and basal cognition, reveals a rich research program with many implications for cognitive science, evolutionary biology, regenerative medicine, and artificial intelligence.
... Moreover, because the same Vmem dynamics can be produced by many different ion channel combinations, and because bioelectric states propagate their influence across tissue distance during morphogenesis [264; 265], evolution is free to swap out channels and explore the bioelectrical state space: simple mutations in electrogenic genes can exert very long-range, highly coordinated changes in anatomy. Indeed, the KCNH8 ion channel and a connexin were identified in the transcriptomic analysis of the evolutionary shift between two functionally different morphologies of fin structures in fish [266]. The evolutionary significance of bioelectric controls can also be seen across lineages, as some viruses evolved to carry ion channel and gap junction (Vinnexin) genes that enable them to hijack bioelectric machinery used by their target cells [267; 268]. ...
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Synthetic biology and bioengineering provide the opportunity to create novel embodied cognitive systems (otherwise known as minds) in a very wide variety of chimeric architectures combining evolved and designed material and software. These advances are disrupting familiar concepts in the philosophy of mind, and require new ways of thinking about and comparing truly diverse intelligences, whose composition and origin are not like any of the available natural model species. In this Perspective, I introduce TAME - Technological Approach to Mind Everywhere - a framework for understanding and manipulating cognition in unconventional substrates. TAME formalizes a non-binary (continuous), empirically-based approach to strongly embodied agency. When applied to regenerating/developmental systems, TAME suggests a perspective on morphogenesis as an example of basal cognition. The deep symmetry between problem-solving in anatomical, physiological, transcriptional, and 3D (traditional behavioral) spaces drives specific hypotheses by which cognitive capacities can scale during evolution. An important medium exploited by evolution for joining active subunits into greater agents is developmental bioelectricity, implemented by pre-neural use of ion channels and gap junctions to scale cell-level feedback loops into anatomical homeostasis. This architecture of multi-scale competency of biological systems has important implications for plasticity of bodies and minds, greatly potentiating evolvability. Considering classical and recent data from the perspectives of computational science, evolutionary biology, and basal cognition, reveals a rich research program with many implications for cognitive science, evolutionary biology, regenerative medicine, and artificial intelligence.
... 2 The sword is a male restricted trait, but female swordtails develop swords like males when treated with testosterone. 3,4 This finding suggests that a potential sexual conflict has been solved by a strict androgen dependency for expression of the phenotype. Females of Xiphophorus hellerii and several other species preferentially associate with males carrying a longer sword over males with shorter swords, which is thought to result in a higher mating success of long-sworded males. ...
... Because immature fish and adult females also develop a sword indistinguishable from the male structure following treatment with androgens 3,4 we generated (3) an RNA-seq dataset from the sword of testosteronetreated adult females; and added (4) our previous dataset from testosterone-induced swords in pre-pubertal juveniles. 3 Small biopsies from the dorsal and ventral fin margin during a timed series of growth and of regeneration and from the hormoneinduced and naturally developed swords from 15-20 individuals were pooled and used for transcriptome sequencing. To exclude genes that are not involved in sword development but have a more general function during natural and hormone induced caudal fin growth or in regeneration, differential expression The swordless Priapella lacandonae is the nearest (sister genus) and medaka, Oryzias latipes, a distant outgroup. ...
Article
Sexual selection results in sex-specific characters like the conspicuously pigmented extension of the ventral tip of the caudal fin—the “sword”—in males of several species of Xiphophorus fishes. To uncover the genetic architecture underlying sword formation and to identify genes that are associated with its development, we characterized the sword transcriptional profile and combined it with genetic mapping approaches. Results showed that the male ornament of swordtails develops from a sexually non-dimorphic prepattern of transcription factors in the caudal fin. Among genes that constitute the exclusive sword transcriptome and are located in the genomic region associated with this trait we identify the potassium channel, Kcnh8, as a sword development gene. In addition to its neural function kcnh8 performs a known role in fin growth. These findings indicate that during evolution of swordtails a brain gene has been co-opted for an additional novel function in establishing a male ornament.
... Hence, the dorsal fin-specific L > S expression in S. casuarius might reflect more active neural regeneration in the distinctly elongated region of the dorsal fin than in the moderately elongated anal fin, although L > S expression at stage 0 also indicates a role in innervation maintenance. Differences in the L/S expression patterns of cx43, mmp9 and foxd3 between the two fin types are particularly interesting because of their proposed regulatory interactions and their roles in fin ray growth 11,16,[18][19][20] . In zebrafish, the decreased expression of cx43 leads to up-regulation of mmp9 in the caudal fin 18 and to defects in the lengthening of bony fin ray segments 19 . ...
... L < S expression) was observed only in the dorsal fin, where the difference in segment length between elongated and short fin rays was more pronounced than in the anal fin. Similarly, the transcription factor foxd3, which was associated with the exaggerated fin outgrowth (the sword) of male sword-tail fish 11 and proposed as a main upstream regulator of the fin elongation GRN in N. brichardi 16 , showed significant L > S expression only in the dorsal fin of S. casuarius. In zebrafish, foxd3 is implicated in the dedifferentiation associated with tissue regeneration through its balancing effects on BMP and Wnt signals 37,38 . ...
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Comparative analyses of gene regulation inform about the molecular basis of phenotypic trait evolution. Here, we address a fin shape phenotype that evolved multiple times independently across teleost fish, including several species within the family Cichlidae. In a previous study, we proposed a gene regulatory network (GRN) involved in the formation and regeneration of conspicuous filamentous elongations adorning the unpaired fins of the Neolamprologus brichardi. Here, we tested the members of this network in the blockhead cichlid, Steatocranus casuarius, which displays conspicuously elongated dorsal and moderately elongated anal fins. Our study provided evidence for differences in the anatomy of fin elongation and suggested gene regulatory divergence between the two cichlid species. Only a subset of the 20 genes tested in S. casuarius showed the qPCR expression patterns predicted from the GRN identified in N. brichardi, and several of the gene-by-gene expression correlations differed between the two cichlid species. In comparison to N. brichardi, gene expression patterns in S. casuarius were in better (but not full) agreement with gene regulatory interactions inferred in zebrafish. Within S. casuarius, the dorsoventral asymmetry in ornament expression was accompanied by differences in gene expression patterns, including potential regulatory differentiation, between the anal and dorsal fin.
... Extensive research has been launched to identify genes underlying fin growth and regeneration with a strong focus on the caudal fin of the zebrafish model Danio rerio 2,[8][9][10] . It is only in recent years that the molecular basis of the morphological diversity of fins within and across species has attracted some attention [11][12][13][14][15] . Studies capitalizing on the natural variation in fin morphology addressed, for instance, the ventral elongation of the caudal fin in swordtail fish 11 , interspecific divergence in pectoral fin morphology in cichlids from Lake Malawi 13 and the twin-tail phenotype of goldfish 14 . ...
... It is only in recent years that the molecular basis of the morphological diversity of fins within and across species has attracted some attention [11][12][13][14][15] . Studies capitalizing on the natural variation in fin morphology addressed, for instance, the ventral elongation of the caudal fin in swordtail fish 11 , interspecific divergence in pectoral fin morphology in cichlids from Lake Malawi 13 and the twin-tail phenotype of goldfish 14 . Here, we are interested in the molecular basis of fin filaments, that is, ornamental elongations of fins which are displayed by numerous fish species across various taxonomic groups. ...
... Some of the genes detected in the present study have already been implicated in studies of teleost fish fin regeneration and morphogenesis (see details in Table 1). These include anxa2a, a member of the annexin family 36 , two angiopoietic protein encoding genes, angptl5 and angptl7 11,37 , dpysl5a, which encodes a member of the Collapsin response mediator protein (CRMP) family 38 , and c1qtnf5, encoding a basement membrane component 39 . Some other members of the gene network are not directly indicated in fin regeneration but appeared to have related functions in vertebrates. ...
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Variation in fin shape and size contributes to the outstanding morphological diversity of teleost fishes, but the regulation of fin growth has not yet been studied extensively outside the zebrafish model. A previous gene expression study addressing the ornamental elongations of unpaired fins in the African cichlid fish Neolamprologus brichardi identified three genes (cx43, mmp9 and sema3d) with strong and consistent expression differences between short and elongated fin regions. Remarkably, the expression patterns of these genes were not consistent with inferences on their regulatory interactions in zebrafish. Here, we identify a gene expression network (GRN) comprising cx43, mmp9, and possibly also sema3d by a stepwise approach of identifying co-expression modules and predicting their upstream regulators. Among the transcription factors (TFs) predicted as potential upstream regulators of 11 coexpressed genes, six TFs (foxc1, foxp1, foxd3, myc, egr2, irf8) showed expression patterns consistent with their cooperative transcriptional regulation of the gene network. Some of these TFs have already been implicated in teleost fish fin regeneration and formation. We particularly discuss the potential function of foxd3 as driver of the network and its role in the unexpected gene expression correlations observed in N. brichardi.
... Hence, little is known about whether these gene regulatory networks determine fin shape traits, such as regional outgrowth, as well. In swordtail fish, where males grow a sword-like elongation on the ventral part of their caudal fin, sword-specific gene expression patterns were detected for a number of genes 14,25,26 . ...
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The diversity of fin morphology within and across fish taxa offers great, but still largely unexplored, opportunities to investigate the proximate mechanisms underlying fin shape variation. Relying on available genetic knowledge brought forth mainly by the comprehensive study of the zebrafish caudal fin, we explored candidate molecular mechanisms for the maintenance and formation of the conspicuously elongated filaments adorning the unpaired fins of the East African "princess cichlid" Neolamprologus brichardi. Via qPCR assays, we detected expression differences of candidate genes between elongated and short regions of intact and regenerating fins. The identified genes include skeletogenic and growth factors (igf2b, fgf3, bmp2 and bmp4), components of the WNT pathway (lef1, wnt5b and wnt10) and a regulatory network determining fin ray segment size and junction (cx43, esco2 and sema3d), as well as other genes with different roles (mmp9, msxb and pea3). Interestingly, some of these genes showed fin specific expression differences which are often neglected in studies of model fish that focus on the caudal fin. Moreover, while the observed expression patterns were generally consistent with zebrafish results, we also detected deviating expression correlations and gene functions.
... In particular, RNA-Seq facilitates the study of the gene regulatory networks of ecologically or evolutionarily intriguing traits in nonmodel organisms, even if their genomes have not been sequenced yet (e.g., Elmer et al. 2010). Several recent studies using RNA-Seq have been performed to study gene expression patterns and genetic pathways underlying key ecological traits in fishes (e.g., Gunter et al. 2013;Henning et al. 2013;Manousaki et al. 2013;Kang et al. 2015). Yet, the genetic underpinnings of adaptive behavioral phenotypes and the role of gene expression differences in regulating behavior remain largely unidentified (e.g., Whitfield et al. 2003;Aubin-Horth et al. 2005;Renn et al. 2008;Drew et al. 2012;Harris and Hofmann 2014). ...
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Lateralized behavior ('handedness') is unusual, but consistently found across diverse animal lineages, including humans. It is thought to reflect brain anatomical and/or functional asymmetries, but its neuro-molecular mechanisms remain largely unknown. Lake Tanganyika scale-eating cichlid fish, Perissodus microlepis show pronounced asymmetry in their jaw morphology as well as handedness in feeding behavior - feeding scales preferentially only from one or the other side of their victims. This makes them an ideal model in which to investigate potential laterality in neuroanatomy and transcription in the brain in relation to behavioral handedness. After determining behavioral handedness in P. microlepis (preferred attack side), we estimated the volume of the hemispheres of brain regions and captured their gene expression profiles. Our analyses revealed that the degree of behavioral handedness is mirrored at the level of neuroanatomical asymmetry, particularly in the tectum opticum. Transcriptome analyses showed that different brain regions (tectum opticum, telencephalon, hypothalamus and cerebellum) display distinct expression patterns, potentially reflecting their developmental interrelationships. For numerous genes in each brain region, their extent of expression differences between hemispheres was found to be correlated with the degree of behavioral lateralization. Interestingly, the tectum opticum and telencephalon showed divergent biases on the direction of up- or down-regulation of the laterality candidate genes (e.g., grm2 ) in the hemispheres, highlighting the connection of handedness with gene expression profiles and the different roles of these brain regions. Hence, handedness in predation behavior may be caused by asymmetric size of brain hemispheres and also by lateralized gene expressions in the brain.
... Thus, SSTs typically evolved in male individuals [1][2][3]. The best studied SSTs are elaborate ornaments or weapons, such as the peacock's tail [4], horns of scarab beetles [5], swords of the swordtail fish [6], and antlers of the deer [7]. Among SSTs, weapons have evolved multiple times across the animal kingdom [8]. ...
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Background The expression of sexually selected traits often varies with populations’ breeding cycles in many animals. The elucidation of mechanisms underlying the expression of such traits is a research topic in evolutionary biology; however, the genetic basis of the seasonal development of their expression remains unknown. Male Leptobrachium boringii develop keratinized nuptial spines on their upper jaw during the breeding season that fall off when the breeding season ends. To illuminate the genetic basis for the expression of this trait and its seasonal development, we assessed the de novo transcriptome for L. boringii using brain, testis and upper jaw skin and compared gene expression profiles of these tissues between two critical periods of the spine growth cycle. ResultsWe identified 94,900 unigenes in our transcriptome. Among them, 2,131 genes were differentially expressed between the breeding period when the spines developed and the post-breeding period when the spines were sloughed. An increased number of differentially expressed genes (DEGs) were identified in the upper jaw skin compared with the testis and brain. In the upper jaw skin, DEGs were mainly enriched in cytosolic part, peptidase inhibitor activity and peptidase regulator activity based on GO enrichment analysis and in glycolysis/gluconeogenesis, ribosome biogenesis in eukaryotes and retinol metabolism based on KEGG enrichment analysis. In the other two tissues, DEGs were primarily involved in the cell cycle, DNA replication and melatonin production. Specifically, insulin/insulin-like growth factor and sex steroid hormone-related DEGs were identified in the upper jaw skin, indicating . The expression variation of IGF2 and estrogen-related genes may be the main factors regulating the seasonal development of the spines. Conclusions Our study provides a list of potential genes involved in the regulation of seasonal development of nuptial spines in L. boringii. This is the first transcriptome survey of seasonally developed sexually selected traits for non-model amphibian species, and candidate genes provided here may provide valuable information for further studies of L. boringii.