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Bioluminescence is defined as the production of visible light by a living organism [1] . To fulfil its ecological role (i.e. defense, offense, or communication), bioluminescence needs to be finely controlled. Although nervous or hormonal control mechanisms have been demonstrated to occur [1-4] , another type of local control via extraocular photo-perception has been suggested to operate in different marine luminescent taxa. Photoemission and photo-perception association imposed a clear co-evolution between the two light-related processes. Recently, extraocular photo-perception processes were shown or suggested to occur at the level of the bioluminescent production site (i.e. photophore/photocyte) in various marine organisms. Photoreceptor molecular markers, including metazoan prototypical photoreceptor proteins (i.e. opsins), have first been identified in the bioluminescent organ of the cephalopod Euprymna scolopes [5] . In this sepiolid, light produced by symbiotic bacteria is perceived by extraocular opsins located in the tissues of the photophore. The discovery of this extraocular photo-perception highlighted the presence of an unsuspected relationship between extraocular perception of light and bioluminescence. Then, studies revealed opsin expression in the photocytes of actenophore, Mnemiopsis leydii, reinforcing the idea of a local interconnexion between light emission and reception, even in an organism producing its own luminescence [6] . More recently, opsins were specifically demonstrated (i) to be expressed within the photocytes of the ophiuroid Amphiura filiformis [7] , the photophores of the smalleye pygmy shark, Squaliolus aliae [4] , the parapodial luminous gland of the polychaete Tomopteris helgolandica [8] , and (ii) to be expressed and functionally active within the photophores of the shrimp Janicella spinicauda [9] . Finally, a deeper characterization of the link between the two processes was made in the lanternshark Etmopterus spinax, where not only opsin characterization and localization within the photophore are depicted, but also the morpho-functional action and intracellular pathways linking light emission and perception [10] . Except for the lanternshark, control mechanisms linking both photobiological processes remain unknown. Despite, functional coupling between photoreception and photoemission is suggested to be an astonishing example of functional convergent evolution in luminescent organisms, further investigations need to be done to understand and highlight this unexpected phylogenetically widespread mechanisms

Biodiversity in a rapidly changing world Louvain-la-Neuve, 28-29 April 2022 ABSTRACT : Coubris Constance, Mallefet Jérôme (UCLouvain) constance.coubris@uclouvain.be Bioluminescence induction in Amphiura filiformis Bioluminescence is the capability to emit visible light thanks to a biochemical reaction. About 80% of the known luminous species live in the marine environment. Among luminous substrates, coelenterazine is the most widespread luciferin found in at least eight phyla. The broad phylogenetic distribution of this light-emitting molecule led to the hypothesis of its dietary acquisition, so far demonstrated in one cnidarian and one lophogastrid shrimp species. Within Ophiuroidea, the dominant class of luminous Echinoderms, Amphiura filiformis is a brittle star model species known to use coelenterazine as substrate in a luciferin/luciferase reaction. A previous study performed long-term monitoring of A. filiformis luminescent capabilities maintained in captivity under controlled conditions. Our results showed (i) depletion of luminescent capabilities within five months when the ophiuroid was fed with a coelenterazine-free diet and (ii) after 15 months in captivity, A. filiformis showed a quick recovery of luminescent capabilities when fed with coelenterazine supplemented food. The results strongly support for the first time the trophic acquisition of coelenterazine in one brittle star. Work is in progress on A. filiformis to find: (i) does seasonal variations of luminescence capabilities exist; (ii) what is the natural source of coelenterazine in the sea, (iii) the transfer mechanism of coelenterazine from the stomach to the luminous cells, and (iv) how often the brittle star has to refill the stock of luminous substrate.

Bioluminescence is the capability to emit visible light thanks to a biochemical reaction. About 80% of the known luminous species live in the marine environment. Among luminous substrates, coelenterazine is the most widespread luciferin found in at least eight phyla. The broad phylogenetic distribution of this light-emitting molecule led to the hypothesis of its dietary acquisition, so far demonstrated in one cnidarian [1] and one lophogastrid shrimp [2] species. Within Ophiuroidea, the dominant class of luminous Echinoderms, Amphiura filiformis is a brittle star model species known to use coelenterazine as substrate in a luciferin/luciferase reaction. A previous study performed long-term monitoring of A. filiformis luminescent capabilities maintained in captivity under controlled conditions [3] . Our results showed (i) depletion of luminescent capabilities within five months when the ophiuroid was fed with a coelenterazine-free diet and (ii) after 15 months in captivity, A. filiformis showed a quick recovery of luminescent capabilities when fed with coelenterazine supplemented food. The results strongly support for the first time the trophic acquisition of coelenterazine in one brittle star. Current work on A. filiformis aims to go further on different aspects: (i) does seasonal variations of luminescence capabilities exist; (ii) what is the natural source of coelenterazine in the Swedish fjörd, (iii) how often the brittle star has to refill the stock of luminous substrate. To date, A. filiformis luminous capabilities were measured in summer, fall, and winter to evaluate potential seasonal variations. In parallel, analysis of the fjord planktonic communities and biochemical assays confirm the presence of species containing coelenterazine within potential A. filiformis preys. Stomach content DNA-barcoding is currently in progress to identify brittle star diet. This strategy will allow making a match between the brittle star's prey and plankton species known to contain coelenterazine. Moreover, long-term monitoring of the brittle star luminous capabilities is performed to evaluate (i) the depletion and reacquisition timing for each season, and (ii) the frequency of the coelenterazine input in the A. filiformis diet to conserve its bioluminescence capabilities.

Luminous sharks are represented in three families, Etmopteridae, Dalatiidae and Somniosidae. Even structural differences in light organs (i.e. photophores) exist between these shark families, they are small and found in great number mainly on the ventral side. Dalatiidae and Somniosidae harbor photophores constituted of a single photocyte (i.e. light emitting cell) encapsulated in a pigmented layer, and topped by one or several lens cells. Etmopteridae photophores are more complex with many photocytes, cells forming an iris-like structure (ILS), as well as a reflector encapsulated within the pigmented layer. Conversely to luminescent bony fishes, shark luminescence is under a hormonal control. Recently, an extraocular opsin has been shown to be involved in the mechanical luminescence regulation through the pigment motion within the ILS. In the shark model, Etmopterus spinax, blue light (i.e. mimicking the shark bioluminescence) absorption has been shown to modify the IP3 concentration within photophores and trigger pathway regulating the ILS cells pigment motions. By analyzing the photophore ontogeny of two luminous sharks (E. spinax for the Etmopteridae and Squaliolus aliae for the Dalatiidae), a similar photophore morphogenesis was highlighted. In parallel, the extraocular opsin, encephalopsin (i.e. Opn3), expression was followed during the photophore development through immunohistology. Opn3 immunoreactivity appeared concomitantly with the photophore final formation in both species, assuming an implication in shark luminescence perception. The similarity between the photophore morphogenesis, opsin expression site and timing of expression suggest that the mechanism involving the Opn3 in the bioluminescence regulation is conserved across luminous shark families during the evolution.

Bioluminescence — the emission of visible light by living organisms — relies on the oxidation of a luciferin substrate catalysed by a luciferase enzyme ([1], for review). This “famous” terminology may give the false idea that all luminous organisms are using identical or homologous molecular tools to achieve bioluminescence. Instead, multiple light emission systems co‐emerged independently along the tree of life resulting in a plethora of non‐homologous luciferases ([2], for review). To investigate the diversity of known luciferase proteins (including luciferases stricto sensu and photoproteins), we analysed a total of 134 sequences coming from 75 species and 11 phyla. The global dataset was analysed using a sequence‐similarity‐based clustering approach based on BLASTp e‐values and using the CLANS software [3]. We identified 12 distinct groups of luciferases, each comprising homologous proteins but with no homology between groups [4]. In addition, non‐luciferase ‐ but homologous – enzymes were added to the analyses to illustrate the evolutionary context of each identified group. It appears that phylogenetically related organisms may sometimes use non‐homologous luciferases (e.g., case of luminous decapods, ostracods, and copepods within Pancrustacea) but also that phylogenetically distant organisms may use homologous luciferases (e.g., case of the brittle star Amphiura filiformis and the sea pansy Renilla). Genes coding for luciferases may then have emerged as new genes or have been co‐opted from ancestral non‐luciferase genes (i.e., with an ancestral function unrelated to bioluminescence). Evolution (and natural selection, in particular) indeed often promotes evolutionary innovation by co‐opting preexisting genes for new functions. In this latter case, the homologous gene co‐ options may occur independently in phylogenetically distant organisms (i.e., parallel evolution) as observed in the case of the brittle star/sea pansy luciferases [5]. As already suggested for other types of proteins (e.g., [6]), our findings suggest that co‐option may be an underappreciated process underpinning protein “neofunctionalisation”, in particular in the context of luciferase evolution.

Despite the reasonable number of luminous species, echinoderm luminescence is still poorly documented [1-2]. In this plenary, recent discoveries about echinoderms luminescence will be presented underlining what are new avenues for future research aiming to understand: (i) how many species of echinoderms glow; (ii) how echinoderms emit light; (iii) how can eyeless echinoderms perceive their light and finally (iv) why are echinoderms able to produce light in the abyss? In Echinoderms, the first luminous compounds isolated by Shimomura in 1986 from a brittle star, Ophiopsila californica, was a photoprotein termed Ophiopsilin [3]. Fifteen years later, the first luciferin/luciferase system based on coelenterazine was reported in Amphiura filiformis [4-5]. Recent studies demonstrated that A. filiformis used a dietary acquisition of coelenterazine to maintain its luminous capability[5]. Works are in progress to understand this diet transfer to restock luminous substrate. Echinoderms have often been viewed to have poor visual abilities due to a lack of well-developed eyes. A. filiformis has also been used to demonstrate the existence of extraocular opsins using transcriptomic methods and it is now well accepted that brittle stars perceive and respond to light stimuli. New research will look at the possible interactions between photoemission and photoreception in this model species[7-8]. In 1977 a first study done on deep- sea echinoderms bioluminescence was produced [9]. Forty years later, “Sampling the Abyss”, an expedition on the East coasts of Australia allowed systematic surveys of abyssal plains from Hobart to the Great Barrier Reef using bottom beam trawls. These brought back at the surface specimens allowing us to document for the first time some bioluminescent ability of rarely seen organisms using a sensitive camera, video recordings, and luminometer. Images and data from benthic abyssal echinoderms belonging to Asteroidea, Ophiuroidea, Crinoidea, and Holothuroidea evealed the diversity of bioluminescence, ranging from slow glow to brief flash, from blue to green emitters[10]. Deep-down spiny critters shine light but for what purpose, this remains a mystery. News collaborations and new expeditions will help to better understand echinoderm’s bioluminescence