The Nereid on the rise: Platynereis as a model system

The Nereid Platynereis dumerilii (Audouin and Milne Edwards (Annales des Sciences Naturelles 1:195–269, 1833) is a marine annelid that belongs to the Nereididae, a family of errant polychaete worms. The Nereid shows a pelago-benthic life cycle: as a general characteristic for the superphylum of Lophotrochozoa/Spiralia, it has spirally cleaving embryos developing into swimming trochophore larvae. The larvae then metamorphose into benthic worms living in self-spun tubes on macroalgae. Platynereis is used as a model for genetics, regeneration, reproduction biology, development, evolution, chronobiology, neurobiology, ecology, ecotoxicology, and most recently also for connectomics and single-cell genomics. Research on the Nereid started with studies on eye development and spiralian embryogenesis in the nineteenth and early twentieth centuries. Transitioning into the molecular era, Platynereis research focused on posterior growth and regeneration, neuroendocrinology, circadian and lunar cycles, fertilization, and oocyte maturation. Other work covered segmentation, photoreceptors and other sensory cells, nephridia, and population dynamics. Most recently, the unique advantages of the Nereid young worm for whole-body volume electron microscopy and single-cell sequencing became apparent, enabling the tracing of all neurons in its rope-ladder-like central nervous system, and the construction of multimodal cellular atlases. Here, we provide an overview of current topics and methodologies for P. dumerilii, with the aim of stimulating further interest into our unique model and expanding the active and vibrant Platynereis community.

Maternal Transcripts of Hox Genes Are Found in Oocytes of Platynereis dumerilii (Annelida, Nereididae)

Hox genes are some of the best studied developmental control genes. In the overwhelming majority of bilateral animals, these genes are sequentially activated along the main body axis during the establishment of the ground plane, i.e., at the moment of gastrulation. Their activation is necessary for the correct differentiation of cell lines, but at the same time it reduces the level of stemness. That is why the chromatin of Hox loci in the pre-gastrulating embryo is in a bivalent state. It carries both repressive and permissive epigenetic markers at H3 histone residues, leading to transcriptional repression. There is a paradox that maternal RNAs, and in some cases the proteins of the Hox genes, are present in oocytes and preimplantation embryos in mammals. Their functions should be different from the zygotic ones and have not been studied to date. Our object is the errant annelid Platynereis dumerilii. This model is convenient for studying new functions and mechanisms of regulation of Hox genes, because it is incomparably simpler than laboratory vertebrates. Using a standard RT-PCR on cDNA template which was obtained by reverse transcription using random primers, we found that maternal transcripts of almost all Hox genes are present in unfertilized oocytes of worm. We assessed the localization of these transcripts using WMISH.

DNA methylation atlas and machinery in the developing and regenerating annelid Platynereis dumerilii

Background Methylation of cytosines in DNA (5mC methylation) is a major epigenetic modification that modulates gene expression and constitutes the basis for mechanisms regulating multiple aspects of embryonic development and cell reprogramming in vertebrates. In mammals, 5mC methylation of promoter regions is linked to transcriptional repression. Transcription regulation by 5mC methylation notably involves the nucleosome remodeling and deacetylase complex (NuRD complex) which bridges DNA methylation and histone modifications. However, less is known about regulatory mechanisms involving 5mC methylation and their function in non-vertebrate animals. In this paper, we study 5mC methylation in the marine annelid worm Platynereis dumerilii, an emerging evolutionary and developmental biology model capable of regenerating the posterior part of its body post-amputation. Results Using in silico and experimental approaches, we show that P. dumerilii displays a high level of DNA methylation comparable to that of mammalian somatic cells. 5mC methylation in P. dumerilii is dynamic along the life cycle of the animal and markedly decreases at the transition between larval to post-larval stages. We identify a full repertoire of mainly single-copy genes encoding the machinery associated with 5mC methylation or members of the NuRD complex in P. dumerilii and show that this repertoire is close to the one inferred for the last common ancestor of bilaterians. These genes are dynamically expressed during P. dumerilii development and regeneration. Treatment with the DNA hypomethylating agent Decitabine impairs P. dumerilii larval development and regeneration and has long-term effects on post-regenerative growth. Conclusions Our data reveal high levels of 5mC methylation in the annelid P. dumerilii, highlighting that this feature is not specific to vertebrates in the bilaterian clade. Analysis of DNA methylation levels and machinery gene expression during development and regeneration, as well as the use of a chemical inhibitor of DNA methylation, suggest an involvement of 5mC methylation in P. dumerilii development and regeneration. We also present data indicating that P. dumerilii constitutes a promising model to study biological roles and mechanisms of DNA methylation in non-vertebrate bilaterians and to provide new knowledge about evolution of the functions of this key epigenetic modification in bilaterian animals.

Jaws of Platynereis dumerilii: Miniature Biogenic Structures with Hardness Properties Similar to Those of Crystalline Metals

Nanoindentation, laser ablation inductively coupled plasma mass spectroscopy and weighing ion-spiked organic matrix standards revealed structure-property relations in the microscopic jaw structures of a cosmopolitan bristle worm, Platynereis dumerilii. Hardness and elasticity values in the jaws’ tip region, exceeding those in the center region, can be traced back to more metal and halogen ions built into the structural protein matrix. Still, structure size appears as an even more relevant factor governing the hardness values measured on bristle worm jaws across the genera Platynereis, Glycera and Nereis. The square of the hardness scales with the inverse of the indentation depth, indicating a Nix-Gao size effect as known for crystalline metals. The limit hardness for the indentation depth going to infinity, amounting to 0.53 GPa, appears to be an invariant material property of the ion-spiked structural proteins likely used by all types of bristle worms. Such a metal-like biogenic material is a major source of bio-inspiration.

Segment number threshold determines juvenile onset of germline cluster proliferation in Platynereis dumerilii

Many animals rely on sexual reproduction to propagate by using gametes (oocytes and sperm). Development of sexual characters and generation of gametes are tightly coupled with the growth of an organism. Platynereis dumerilii is a marine segmented worm which has been used to study germline development and gametogenesis. Platynereis has 4 Primordial Germ Cells (PGCs) that arise early in development and these cells are thought to give rise to germ cell clusters found across the body in the juvenile worms. The germ cell clusters eventually form the gametes. The stages of germline development and how the 4 PGCs become the numerous germ cell clusters are not well-documented in the juvenile stages. Platynereis, like other segmented worms, grows by adding new segments at its posterior end. The number of segments generally reflect the growth state of the worms and therefore is a useful and easily measurable growth state metric. To understand how growth correlates with development and gametogenesis, we investigated germline development across several developmental stages using germline/multipotency markers. We found that segment number predicted the state of germline development and the abundance of germline clusters. Additionally, we found that keeping worms short in segment number via changing external conditions or via amputations supported segment number threshold requirement for germline development. Finally, we asked if these clusters in Platynereis play a role in regeneration (as similar free-roaming cells are observed in Hydra and planarian regeneration) and found that the clusters were not required for regeneration in Platynereis, suggesting a strictly germline nature. Overall, these molecular analyses suggest a previously unidentified developmental transition dependent on the growth state in juvenile Platynereis when germline proliferation is substantially increased.

Two light sensors decode moonlight versus sunlight to adjust a plastic circadian/circalunidian clock to moon phase

Many species synchronize their physiology and behavior to specific hours. It is commonly assumed that sunlight acts as the main entrainment signal for ~24h clocks. However, the moon provides similarly regular time information, and increasingly studies report correlations between diel behavior and lunidian cycles. Yet, mechanistic insight into the possible influences of the moon on ~24hr timers is scarce. We studied Platynereis dumerilii and uncover that the moon, besides its role in monthly timing, also schedules the exact hour of nocturnal swarming onset to the nights′ darkest times. Moonlight adjusts a plastic clock, exhibiting <24h (moonlit) or >24h (no moon) periodicity. Abundance, light sensitivity, and genetic requirement indicate Platynereis r-Opsin1 as receptor to determine moonrise, while the cryptochrome L-Cry is required to discriminate between moon- and sunlight valence. Comparative experiments in Drosophila suggest that Cryptochrome′s requirement for light valence interpretation is conserved. Its exact biochemical properties differ, however, between species with dissimilar timing ecology. Our work advances the molecular understanding of lunar impact on fundamental rhythmic processes, including those of marine mass spawners endangered by anthropogenic change.

A Cryptochrome adopts distinct moon- and sunlight states and functions as moonlight interpreter in monthly oscillator entrainment

Measuring time by the moon's monthly cycles is a wide-spread phenomenon and crucial for successful reproduction in countless marine organisms. In several species, such as the mass-spawning bristle worm Platynereis dumerilii, an endogenous monthly oscillator synchronizes reproduction to specific days. Classical work showed that this oscillator is set by full moon. But how do organisms recognize this specific moon phase? We uncover L-Cry's involvement: photoreduction and recovery kinetics of its co-factor FAD differ strongly when purified L-Cry is exposed to naturalistic moonlight, naturalistic sunlight, or their different successions. L-Cry's sun- versus moonlight states correlate with distinct sub-cellular localizations, indicating differential signalling. These properties enable a discrimination between sun- and moonlight, as well as moonlight duration as a moon phase indicator. Consistently, l-cry mutants re-entrain their circalunar phase less well than wild-type to naturalistic moonlight. But under artificially strong nocturnal light, l-cry mutants re-entrain faster than wildtype, suggesting that L-Cry at least partly blocks "wrong" light from impacting on this oscillator. Our work provides a new level of functional understanding of moon-regulated biological processes.

The small polychaete Platynereis dumerilii revealed as a large species complex with fourteen MOTUs in European marine habitats

Recent studies reporting complexes of cryptic or pseudo-cryptic species with narrow geographic distributions have been challenging the cosmopolitan status of a fair number of marine benthic invertebrates. Morphologically similar species are often overlooked but molecular techniques have been extremely effective in signalling potential hidden diversity which, complemented with further detailed examination, might reveal unique morphological and ecological features. Evidence of morphological stasis, where no clear and stable morphological differences are apparent, can be exemplified by the annelids Platynereis dumerilii (Audouin &amp; Milne Edwards, 1833) and Platynereis massiliensis (Moquin-Tandon, 1869). These sibling species, usually found among algae in marine intertidal and subtidal habitats, can only be distinguished by their different reproductive strategies and life histories. The former is gonochoric, with a single reproductive event in life (semelparous) transforming into a pelagic epitokous form called heteronereis, has free spawning synchronized by lunar periodicity and a larval stage with planktotrophic development; while the latter shows no epitokous transformation and is a protandrous hermaphrodite, characterized by egg brooding and lecithotrophic larval stages with a semi-direct development. In order to verify the possible existence of additional hidden Platynereis species within the P. dumerilii morphotype, we used a multi-locus approach to investigate 26 populations along Europe from the NE Atlantic and the Macaronesia islands (Azores, Madeira and Canaries) to the Western and Eastern Mediterranean Sea. We concatenated the mtDNA COI-5P, rDNA 16S and 28S-D2 sequences and performed a phylogenetic analysis through Bayesian inference (BI). To depict Molecular Operational Taxonomic Units (MOTUs), we applied three delimitation methods (ABGD, bPTP and GYMC) to the concatenated alignment, except for COI where we also applied the Barcode Index Number (BIN), implemented in BOLD, which is exclusive to this locus. Consensus MOTUs were defined based on the majority rule and, in case of draw, the most conservative MOTUs were chosen. We detected at least 14 MOTUs with 23.1% COI mean K2P distance (6.6 - 32.6%). The BI tree is split into three major clades (Clade A: MOTUs 1-3, Clade B: MOTUs 4-9 and Clade C: MOTUs 10-14), with MOTU 3 appearing to represent P. dumerilii sensu stricto and MOTU 9 P. massiliensis. This assumption is based on data from the Type locality and a previous study by Wage et al. (2017), which combined phylogeographic (COI barcode region), reproductive biology and life-history observations on some selected Platynereis populations thriving in the vent areas from the Italian islands of Ischia and Vulcano. Major Cade C seems to be pseudo-cryptic as some visible differences can be found in the specimens, as for example, the size of the tentacular cirri and lack of dorsal pigmentation, in contrast with Clades A and B, where only differences in pigmentation types were found so far. The Spanish archipelago of the Canary Islands and the whole Mediterranean Sea seem to be a cryptic hotspot. Five MOTUs are unique to the Macaronesia and five sympatric MOTUs are present in the Gran Canaria and La Palma islands alone. Additionally, three lineages were present exclusively in the Mediterranean with four sympatric MOTUs spotted in the southeast of Spain (Calpe) and the Greek island of Crete. Three out of four NE Atlantic MOTUs are shared with the Mediterranean with one exclusive to this part of the continent. Failure to recognise this hidden biodiversity may compromise the accuracy and the interpretation of biomonitoring data or other relevant ecological studies. Integrative taxonomy is thus essential to solve these uncertainties and to allow naming the involved undescribed species. Otherwise, most molecular data providing enough support for species hypothesis will continue to be unused, and large fractions of biodiversity will persist unnoticed.

Small-molecule mimicry hunting strategy in the imperial cone snail, Conus imperialis

Venomous animals hunt using bioactive peptides, but relatively little is known about venom small molecules and the resulting complex hunting behaviors. Here, we explored the specialized metabolites from the venom of the worm-hunting cone snail, Conus imperialis. Using the model polychaete worm Platynereis dumerilii, we demonstrate that C. imperialis venom contains small molecules that mimic natural polychaete mating pheromones, evoking the mating phenotype in worms. The specialized metabolites from different cone snails are species-specific and structurally diverse, suggesting that the cones may adopt many different prey-hunting strategies enabled by small molecules. Predators sometimes attract prey using the prey’s own pheromones, in a strategy known as aggressive mimicry. Instead, C. imperialis uses metabolically stable mimics of those pheromones, indicating that, in biological mimicry, even the molecules themselves may be disguised, providing a twist on fake news in chemical ecology.