The mTOR pathway controls the balance between anabolic and catabolic activities in animal cells, acting as a key coordinator of metabolic homeostasis. In fact, the activation of this conserved regulatory circuit promotes the biosynthesis of different macromolecules, including proteins, lipids and nucleic acids, and at the same time blocks catabolic processes such as lysosome biogenesis. In this work, we describe a biological system in which these two aspects of the mTOR function are uncoupled. Studying the sea urchin Paracentrotus lividus, we have found that the activation of the mTOR pathway in the fertilised egg, as well as stimulating protein synthesis, contributes to the development of a dense array of acidic vesicles that behave as lysosomes. We present evidence indicating that mTOR could operate in this context enhancing the translation of the maternal transcripts that code for the multiple components of these organelles. We argue that the mTOR-mediated implementation of a typical catabolic process may in fact support the biosynthetic vocation of this pathway, providing energy and recycled blocks for construction.

The study of early development in sea urchins has been instrumental in uncovering the role of mitotic cyclins, but we still know little about how the specific translation of the mRNAs encoding these proteins impacts mitotic division dynamics. Here, we demonstrate for the first time, in the sea urchin Sphaerechinus granularis (Lamarck, 1816), that cyclin A mRNA is recruited into polysomal fractions following fertilization. Morpholino microinjection targeting cyclin A and B mRNAs delays but does not completely block the first embryonic cleavage. Our results suggest that the translation of Cyclin A and B mRNAs contributes to the fine-tuning of post-fertilization mitotic dynamics, but is not sufficient on its own to achieve the complete process of the first division of sea urchin embryos. This study highlights the importance of the regulation of mitotic Cyclin mRNAs translation in early embryonic development and suggests that the translation of mRNAs encoding additional factors is required for the successful of cell division.

Sea urchins are emblematic models in developmental biology and display several characteristics that set them apart from other deuterostomes. To uncover the genomic cues that may underlie these specificities, we generated a chromosome-scale genome assembly for the sea urchin Paracentrotus lividus and an extensive gene expression and epigenetic profiles of its embryonic development. We found that, unlike vertebrates, sea urchins retained ancestral chromosomal linkages but underwent very fast intrachromosomal gene order mixing. We identified a burst of gene duplication in the echinoid lineage and showed that some of these expanded genes have been recruited in novel structures (water vascular system, Aristotle’s lantern, and skeletogenic micromere lineage). Finally, we identified gene-regulatory modules conserved between sea urchins and chordates. Our results suggest that gene-regulatory networks controlling development can be conserved despite extensive gene order rearrangement.

In the context of the 150th anniversary of the biological station of Roscoff, we describe here how cellular and molecular biology using marine metazoans has been developed at the station during the last 50 years. The discipline was first initiated by an interest in the endocrinology of marine invertebrates and allowed the characterization of the famous Maturation-Promoting Factor (MPF) in echinoderms. During this exciting period of time the biological station focused the interest of the international cell cycle scientific community. Then, discoveries made at the station played a founding role in developing applied research and creating companies offering innovative therapeutic solutions. Later, the scientific dynamics using marine metazoans has been perpetuated by studying the cellular and molecular mechanisms, such as ionic membrane transports or mRNA translation, that govern the life of cells and embryos. More recently, acquiring metazoan genomes has allowed researchers to analyze marine metazoan embryonic development in an evolutionary context. Finally, the Biological Station contributes to transfer this essential knowledge through international courses, which prepare the young generations for future scientific challenges.

During the first steps of sea urchin development, fertilization elicits a marked increase in protein synthesis essential for subsequent cell divisions. While the translation of mitotic cyclin mRNAs is crucial, we hypothesized that additional mRNAs must be translated to finely regulate the onset into mitosis. One of the maternal mRNAs recruited onto active polysomes at this stage codes for the initiation factor eIF4B. Here, we show that the sea urchin eIF4B orthologs present the four specific domains essential for eIF4B function and that Paracentrotus lividus eIF4B copurifies with eIF4E in a heterologous system. In addition, we investigated the role of eIF4B mRNA de novo translation during the two first embryonic divisions of two species, P. lividus and Sphaerechinus granularis. Our results show that injection of a morpholino directed against eIF4B mRNA results in a downregulation of translational activity and delays cell division in these two echinoids. Conversely, injection of an mRNA encoding for P. lividus eIF4B stimulates translation and significantly accelerates cleavage rates. Taken together, our findings suggest that eIF4B mRNA de novo translation participates in a conserved regulatory loop that contributes to orchestrating protein synthesis and modulates cell division rhythm during early sea urchin development.

Echinoderms, a sister group of chordates, is a group of exclusively marine animals. The Echinodermata is an ancient phylum dating to at least 450 million years old and including more than 10,000 extant species present throughout the world’s oceans. The modern echinoderms belong to five classes: Echinoidea (e.g., sea urchins and sand dollars), Asteroidea (e.g., starfishes), Ophiuroidea (e.g., brittle stars), Holothuroidea (e.g., sea cucumbers) and Crinoidea (e.g., sea lilies and feather stars). Sea urchin represents a well-established marine model in biological sciences. This chapter provides a general description of echinoderms and focuses on the significant advances in cell and developmental biology that the study of sea urchins has made possible. During the last decade, many genomic data concerning echinoderms and sea urchins in particular have become available. These new molecular tools have facilitated gene regulation analysis during development and have boosted the possibilities offered by sea urchins as experimental models.

The sea urchin embryo provides a valuable system to analyse the molecular mechanisms orchestrating cell cycle progression and mitosis in a developmental context. However, although it is known that the regulation of histone activity by post-translational modification plays an important role during cell division, the dynamics and the impact of these modifications have not been characterised in detail in a developing embryo. Using different immuno-detection techniques, we show that the levels of Histone 3 phosphorylation at Threonine 3 oscillate in synchrony with mitosis in Sphaerechinus granularis early embryos. We present, in addition, the results of a pharmacological study aimed at analysing the role of this key histone post-translational modification during sea urchin early development.

A major challenge in medical research resides in controlling the molecular processes of tissue regeneration, as organ and structure damage are central to several human diseases. A survey of the literature reveals that mTOR (mechanistic/mammalian target of rapamycin) is involved in a wide range of regeneration mechanisms in the animal kingdom. More particularly, cellular processes such as growth, proliferation, and differentiation are controlled by mTOR. In addition, autophagy, stem cell maintenance or the newly described intermediate quiescence state, Galert, imply upstream monitoring by the mTOR pathway. In this review, we report the role of mTOR signaling in reparative regenerations in different tissues and body parts (e.g., axon, skeletal muscle, liver, epithelia, appendages, kidney, and whole-body), and highlight how the mTOR kinase can be viewed as a therapeutic target to boost organ repair. Studies in this area have focused on modulating the mTOR pathway in various animal models to elucidate its contribution to regeneration. The diversity of metazoan species used to identify the implication of this pathway might then serve applied medicine (in better understanding what is required for efficient treatments in human diseases) but also evolutionary biology. Indeed, species-specific differences in mTOR modulation can contain the keys to appreciate why certain regeneration processes have been lost or conserved in the animal kingdom.

Protein synthesis is a major regulatory step of gene expression in different physiological processes including development. Translation of proteins in sea urchin is stimulated upon fertilization and is necessary for cell cycle progression and development. Translational control is exerted through multifactorial mechanisms, including mRNA recruitment into polysomes and increased rates of translational activity. In this chapter, we review the methods used in sea urchin eggs and embryos to analyze translation activity in vivo both from perspectives of the proteins and of the mRNAs. First, we describe methods to quantify or visualize newly synthesized proteins with radioactive and non-radioactive labeling techniques. Next we present the polysome isolation and profiling on sucrose gradients, allowing the identification of translated mRNAs. Finally, we outline a procedure to follow the translation of a reporter luciferase protein from an mRNA microinjected into the egg.

Sea urchin early development is a powerful model to study translational regulation under physiological conditions. Fertilization triggers an activation of the translation machinery responsible for the increase of protein synthesis necessary for the completion of the first embryonic cell cycles. The cap-binding protein eIF4E, the helicase eIF4A and the large scaffolding protein eIF4G are assembled upon fertilization to form an initiation complex on mRNAs involved in cap-dependent translation initiation. The presence of these proteins in unfertilized and fertilized eggs has already been demonstrated, however data concerning the translational status of translation factors are still scarce. Using polysome fractionation, we analyzed the impact of fertilization on the recruitment of mRNAs encoding initiation factors. Strikingly, whereas the mRNAs coding eIF4E, eIF4A, and eIF4G were not recruited into polysomes at 1 h post-fertilization, mRNAs for eIF4B and for non-canonical initiation factors such as DAP5, eIF4E2, eIF4E3, or hnRNP Q, are recruited and are differentially sensitive to the activation state of the mechanistic target of rapamycin (mTOR) pathway. We discuss our results suggesting alternative translation initiation in the context of the early development of sea urchins.

Marennine is a water-soluble blue-green pigment produced by the marine diatom Haslea ostrearia. The diatom and its pigment are well known from oyster farming areas as the source of the greening of oyster gills, a natural process increasing their market value in Western France. Blooms of blue Haslea are also present outside oyster ponds and hence marine organisms can be exposed, periodically and locally, to significant amounts of marennine in natural environments. Due to its demonstrated antibacterial activities against marine pathogenic bacteria (e.g. Vibrio) and possible prophylactic effects toward bivalve larvae, marennine is of special interest for the aquaculture industry, especially bivalve hatcheries. The present study aimed to provide new insights into the effects of marennine on a large spectrum of marine organisms belonging to different phyla, including species of aquaculture interest and organisms frequently employed in standardised ecotoxicological assays. Different active solutions containing marennine were tested: partially purified Extracellular Marennine (EMn), and concentrated solutions of marennine present in H. ostrearia culture supernatant; the Blue Water (BW) and a new process called Concentrated Supernatant (CS). Biological effects were meanwhile demonstrated in invertebrate species for the three marennine-based solutions at the highest concentrations tested (e.g., decrease of fertilization success, delay of embryonic developmental stages or larval mortality). Exposure to low concentrations did not impact larval survival or development and even tended to enhance larval physiological state. Furthermore, no effects of marennine were observed on the fish gill cell line tested. Marennine could be viewed as a Jekyll and Hyde molecule, which possibly affects the earliest stages of development of some organisms but with no direct impacts on adults. Our results emphasize the need to determine dosages that optimize beneficial effects and critical concentrations not to be exceeded before considering the use of marennine in bivalve or fish hatcheries.

Early embryogenesis relies on the translational regulation of maternally stored mRNAs. In sea urchin, fertilization triggers a dramatic rise in translation activity, necessary for the onset of cell division. Here, the full spectrum of the mRNAs translated upon fertilization was investigated by polysome profiling and sequencing. The translatome of the early sea urchin embryo gave a complete picture of the polysomal recruitment dynamics following fertilization. Our results indicate that only a subset of maternal mRNAs were selectively recruited onto polysomes, with over-represented functional categories in the translated set. The increase in translation upon fertilization depends on the formation of translation initiation complexes following mTOR pathway activation. Surprisingly, mTOR pathway inhibition differentially affected polysomal recruitment of the newly translated mRNAs, which thus appeared either mTOR-dependent or mTOR-independent. Therefore, our data argue for an alternative to the classical cap-dependent model of translation in early development. The identification of the mRNAs translated following fertilization helped assign translational activation events to specific mRNAs. This translatome is the first step to a comprehensive analysis of the molecular mechanisms governing translation upon fertilization and the translational regulatory networks that control the egg-to-embryo transition as well as the early steps of embryogenesis.

We review here previous theoretical and experimental works, which aim to model major events that occur at the time of fertilization in the sea urchin. We discuss works that perform experiments and develop hypotheses that link different scales of biological systems such as the intracellular Ca concentration oscillations and the swimming behavior of sperm, the Ca wave propagation and the fertilization membrane elevation of the egg, and the mRNA translational activation and the completion of the first mitotic division of the early embryo. The aim of this review is on one hand, to highlight the value of systems biology for understanding the mechanisms associated with fertilization and early embryonic development in sea urchins. On the other hand, this review attempts to illustrate, for mathematicians and bioinformaticians, the potential that represent these molecular and cellular events for modeling clear physiological processes.

The fine spatial and temporal resolution of translation control can have a rapid and subtle effect on the microenvironment of the cell in comparison with transcriptional regulation. Mammalian oocyte represents a relevant model that allows addressing the spatial control of translation during meiotic maturation. The fully-grown mammalian oocyte is transcriptionally quiescent and stored maternal RNAs are used for the completion of meiosis and early embryo development. It was shown three decades ago that during resumption of meiosis, protein synthesis is not necessary for the nuclear envelope breakdown (NEBD) but active protein synthesis is required for the correct formation of the meiotic spindle and progression to metaphase II (1). Therefore mammalian oocyte represents a suited system to address translation control in relation with the cell cycle in general and with the spindle formation in particular. In this issue of the Cell Cycle the study by Jansova et al. gains new insight into the role of the eukaryotic initiation factor 4E-Binding Protein 1 (4E-BP1) in the meiotic spindle formation in mouse oocyte (2).

The fine spatial and temporal resolution of translation control can have a rapid and subtle effect on the microenvironment of the cell in comparison with transcriptional regulation. Mammalian oocyte represents a relevant model that allows addressing the spatial control of translation during meiotic maturation. The fully-grown mammalian oocyte is transcriptionally quiescent and stored maternal RNAs are used for the completion of meiosis and early embryo development. It was shown three decades ago that during resumption of meiosis, protein synthesis is not necessary for the nuclear envelope breakdown (NEBD) but active protein synthesis is required for the correct formation of the meiotic spindle and progression to metaphase II (1). Therefore mammalian oocyte represents a suited system to address translation control in relation with the cell cycle in general and with the spindle formation in particular. In this issue of the Cell Cycle the study by Jansova et al. gains new insight into the role of the eukaryotic initiation factor 4E-Binding Protein 1 (4E-BP1) in the meiotic spindle formation in mouse oocyte (2).

In this work, novel archaeosomes based on Egg-PC and a mixture of PEGylated archaeal tetraether lipids were investigated as nanocarriers forin vitrodelivery of an original anticancer peptide.

The cyclin B/CDK1 complex is a key regulator of mitotic entry. Using PP242, a specific ATP-competitive inhibitor of mTOR kinase, we provide evidence that the mTOR signalling pathway controls cyclin B mRNA translation following fertilization in Sphaerechinus granularis and Paracentrotus lividus. We show that PP242 inhibits the degradation of the cap-dependent translation repressor 4E-BP (eukaryotic initiation factor 4E-Binding Protein). PP242 inhibits global protein synthesis, delays cyclin B accumulation, cyclin B/CDK1 complex activation and consequently entry into the mitotic phase of the cell cycle triggered by fertilization. PP242 inhibits cyclin B mRNA recruitment into active polysomes triggered by fertilization. An amount of cyclin B mRNA present in active polysomes appears to be insensitive to PP242 treatment. Taken together, our results suggest that, following sea urchin egg fertilization, cyclin B mRNA translation is controlled by two independent mechanisms: a PP242-sensitive and an additional PP242-insentitive mechanism.

During the past decade, there has been growing interest in the role of translational regulation of gene expression in many organisms. Polysome profiling has been developed to infer the translational status of a specific mRNA species or to analyze the translatome, i.e. the subset of mRNAs actively translated in a cell. Polysome profiling is especially suitable for emergent model organisms for which genomic data are limited. In this paper, we describe an optimized protocol for the purification of sea urchin polysomes and highlight the critical steps involved in polysome purification. We applied this protocol to obtain experimental results on translational regulation of mRNAs following fertilization. Our protocol should prove useful for integrating the study of the role of translational regulation in gene regulatory networks in any biological model. In addition, we demonstrate how to carry out high-throughput processing of polysome gradient fractions, for the simultaneous screening of multiple biological conditions and large-scale preparation of samples for next-generation sequencing.

SUMMARY Sea urchin eggs exhibit a cap‐dependent increase in protein synthesis within minutes after fertilization. This rise in protein synthesis occurs at a constant rate for a great number of proteins translated from the different available mRNAs. Surprisingly, we found that cyclin B, a major cell‐cycle regulator, follows a synthesis pattern that is distinct from the global protein population, so we developed a mathematical model to analyze this dissimilarity in biosynthesis kinetic patterns. The model includes two pathways for cyclin B mRNA entry into the translational machinery: one from immediately available mRNA (mRNAcyclinB) and one from mRNA activated solely after fertilization (XXmRNAcyclinB). Two coefficients, α and β, were added to fit the measured scales of global protein and cyclin B synthesis, respectively. The model was simplified to identify the synthesis parameters and to allow its simulation. The calculated parameters for activation of the specific cyclin B synthesis pathway after fertilization included a kinetic constant (k a ) of 0.024 sec −1 , for the activation of XXmRNAcyclinB, and a critical time interval (t 2 ) of 42 min. The proportion of XXmRNAcyclinB form was also calculated to be largely dominant over the mRNAcyclinB form. Regulation of cyclin B biosynthesis is an example of a select protein whose translation is controlled by pathways that are distinct from housekeeping proteins, even though both involve the same cap‐dependent initiation pathway. Therefore, this model should help provide insight to the signaling utilized for the biosynthesis of cyclin B and other select proteins. Mol. Reprod. Dev. 83: 1070–1082, 2016. © 2016 Wiley Periodicals, Inc .