eIF4B mRNA Translation Contributes to Cleavage Dynamics in Early Sea Urchin Embryos.

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.

Digital Marine: An online platform for blended learning in a marine experimental biology module, the Schmid Training Course.

For over 20 years, the Schmid Training Course (STC) has offered unique opportunities for marine biology students from European universities to learn about marine model organisms. While the topics of the course have continuously changed over the years with the advent of new research techniques and discoveries, the pedagogical approach has remained largely the same - a combination of lectures, lab practicals, and field excursions. Several life science researchers, who have taught in the STC for many years, sought to bring the course's pedagogical approach into the 21st century, and with the support of Erasmus+ Programme of the European Community funding, the Digital Marine project was developed. Digital Marine began in 2018 as an international partnership between the six research centers from which the STC instructors hail, and its main objective was to introduce a flipped, blended approach to learning and teaching with respect to established and emerging marine biological model systems. The Digital Marine platform, which covers 12 marine model organisms, is now publicly available.

mTOR Signaling at the Crossroad between Metazoan Regeneration and Human Diseases.

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.

Genetic and Molecular Insights Into Genotype-Phenotype Relationships in Osteopathia Striata With Cranial Sclerosis (OSCS) Through the Analysis of Novel Mouse Wtx Mutant Alleles.

The X-linked WTX/AMER1 protein constitutes an important component of the ß-catenin destruction complex that can both enhance and suppress canonical ß-catenin signaling. Somatic mutations in WTX/AMER1 have been found in a proportion of the pediatric kidney cancer Wilms' tumor. By contrast, germline mutations cause the severe sclerosing bone dysplasia osteopathia striata congenita with cranial sclerosis (OSCS), a condition usually associated with fetal or perinatal lethality in male patients. Here we address the developmental and molecular function of WTX by generating two novel mouse alleles. We show that in addition to the previously reported skeletal abnormalities, loss of Wtx causes severe midline fusion defects including cleft palate and ectopic synostosis at the base of the skull. By contrast, deletion of the C-terminal part of the protein results in only mild developmental abnormalities permitting survival beyond birth. Adult analysis, however, revealed skeletal defects including changed skull morphology and an increased whole-body bone density, resembling a subgroup of male patients carrying a milder, survivable phenotype. Molecular analysis in vitro showed that while ß-catenin fails to co-immunoprecipitate with the truncated protein, partial recruitment appears to be achieved in an indirect manner using AXIN/AXIN2 as a molecular bridge. Taken together our analysis provides a novel model for WTX-caused bone diseases and explains on the molecular level how truncation mutations in this gene may retain some of WTX-protein functions. © 2018 American Society for Bone and Mineral Research.

The evolution of asymmetric photosensitive structures in metazoans and the Nodal connection.

Asymmetries are observed in a great number of taxa in metazoans. More particularly, functional lateralization and neuroanatomical asymmetries within the central nervous system have been a matter of intense research for at least two hundred years. While asymmetries of some paired structures/organs (e.g. eyes, ears, kidneys, legs, arms) constitute random deviations from a pure bilateral symmetry, brain asymmetries such as those observed in the cortex and epithalamus are directional. This means that molecular and anatomical features located on one side of a given structure are observed in most individuals. For instance, in humans, the neuronal tract connecting the language areas is enlarged in the left hemisphere. When asymmetries are fixed, their molecular mechanisms can be studied using mutants displaying different phenotypes: left or right isomerism of the structure, reversed asymmetry or random asymmetry. Our understanding of asymmetry in the nervous system has been widely enriched thanks to the characterization of mutants affecting epithalamus asymmetry. Furthermore, two decades ago, pioneering studies revealed that a specific morphogen, Nodal, active only on one side of the embryo during development is an important molecule in asymmetry patterning. In this review, I have gathered important data bringing insight into the origin and evolution of epithalamus asymmetry and the role of Nodal in metazoans. After a short introduction on brain asymmetries (chapter I), I secondly focus on the molecular and anatomical characteristics of the epithalamus in vertebrates and explore some functional aspects such as its photosensitive ability related to the pineal complex (chapter II). Third, I discuss homology relationship of the parapineal organ among vertebrates (chapter III). Fourth, I discuss the possible origin of the epithalamus, presenting cells displaying photosensitive properties and/or asymmetry in the anterior part of the body in non-vertebrates (chapter IV). Finally, I report Nodal signaling expression data and functional experiments performed in different metazoan groups (chapter V).

A knock-in mouse line conditionally expressing the tumor suppressor WTX/AMER1.

WTX/AMER1 is an important developmental regulator, mutations in which have been identified in a proportion of patients suffering from the renal neoplasm Wilms' tumor and in the bone malformation syndrome Osteopathia Striata with Cranial Sclerosis (OSCS). Its cellular functions appear complex and the protein can be found at the membrane, within the cytoplasm and the nucleus. To understand its developmental and cellular function an allelic series for Wtx in the mouse is crucial. Whereas mice carrying a conditional knock out allele for Wtx have been previously reported, a gain-of-function mouse model that would allow studying the molecular, cellular and developmental role of Wtx is still missing. Here we describe the generation of a novel mouse strain that permits the conditional activation of WTX expression. Wtx fused to GFP was introduced downstream a stop cassette flanked by loxP sites into the Rosa26 locus by gene targeting. Ectopic WTX expression is reported after crosses with several Cre transgenic mice in different embryonic tissues. Further, functionality of the fusion protein was demonstrated in the context of a Wtx null allele.

Slow cycling cells in the continuous dental lamina of Scyliorhinus canicula: new evidence for stem cells in sharks.

In the lesser spotted catshark (Scyliorhinus canicula), as in most non-mammalian vertebrates, the dentition renews throughout life. To contribute to our understanding of how continuous tooth replacement is achieved, we searched for evidence for the presence of stem cells in this species. Three-dimensional reconstructions of juvenile (2-3 weeks post-hatch) specimens showed that tooth families merge imperceptibly with so-called interdental zones within a continuous and permanent dental lamina. Interdental regions are composed of three layers, continuous with cervical loop, middle, and outer dental epithelium of the tooth families, respectively. A BrdU pulse-chase experiment revealed that cell proliferation is initiated in the lingual part of the dental lamina and the resulting population shifts one tooth position towards the oral epithelium in around four to five weeks. In the longest chase time (114 days) label-retaining and arguably non-differentiated cells were present at the lingual border of the dental lamina. These were found in the outer and middle dental epithelium, both within and between tooth families. This area of the dental lamina did not show expression or distribution of Sox2. Our data support the hypothesis that stem cells reside at the lingual border of the continuous dental lamina, more specifically in the middle dental epithelium at the level of the tooth families, and in its extension between the tooth families. To demonstrate their true stemness and their role in continuous tooth replacement, it remains to be shown that these cells have the potential to give rise to a complete new successor.

Snail1-induced partial epithelial-to-mesenchymal transition drives renal fibrosis in mice and can be targeted to reverse established disease.

Progressive kidney fibrosis contributes greatly to end-stage renal failure, and no specific treatment is available to preserve organ function. During renal fibrosis, myofibroblasts accumulate in the interstitium of the kidney, leading to massive deposition of extracellular matrix and organ dysfunction. The origin of myofibroblasts is manifold, but the contribution of an epithelial-to-mesenchymal transition (EMT) undergone by renal epithelial cells during kidney fibrosis is still debated. We show that the reactivation of Snai1 (encoding snail family zinc finger 1, known as Snail1) in mouse renal epithelial cells is required for the development of fibrosis in the kidney. Damage-mediated Snail1 reactivation induces a partial EMT in tubular epithelial cells that, without directly contributing to the myofibroblast population, relays signals to the interstitium to promote myofibroblast differentiation and fibrogenesis and to sustain inflammation. We also show that Snail1-induced fibrosis can be reversed in vivo and that obstructive nephropathy can be therapeutically ameliorated in mice by targeting Snail1 expression. These results reconcile conflicting data on the role of the EMT in renal fibrosis and provide avenues for the design of novel anti-fibrotic therapies.

The ancestral role of nodal signalling in breaking L/R symmetry in the vertebrate forebrain.

Left-right asymmetries in the epithalamic region of the brain are widespread across vertebrates, but their magnitude and laterality varies among species. Whether these differences reflect independent origins of forebrain asymmetries or taxa-specific diversifications of an ancient vertebrate feature remains unknown. Here we show that the catshark Scyliorhinus canicula and the lampreys Petromyzon marinus and Lampetra planeri exhibit conserved molecular asymmetries between the left and right developing habenulae. Long-term pharmacological treatments in these species show that nodal signalling is essential to their generation, rather than their directionality as in teleosts. Moreover, in contrast to zebrafish, habenular left-right differences are observed in the absence of overt asymmetry of the adjacent pineal field. These data support an ancient origin of epithalamic asymmetry, and suggest that a nodal-dependent asymmetry programme operated in the forebrain of ancestral vertebrates before evolving into a variable trait in bony fish.

Mechanisms of endoderm formation in a cartilaginous fish reveal ancestral and homoplastic traits in jawed vertebrates.

In order to gain insight into the impact of yolk increase on endoderm development, we have analyzed the mechanisms of endoderm formation in the catshark S. canicula, a species exhibiting telolecithal eggs and a distinct yolk sac. We show that in this species, endoderm markers are expressed in two distinct tissues, the deep mesenchyme, a mesenchymal population of deep blastomeres lying beneath the epithelial-like superficial layer, already specified at early blastula stages, and the involuting mesendoderm layer, which appears at the blastoderm posterior margin at the onset of gastrulation. Formation of the deep mesenchyme involves cell internalizations from the superficial layer prior to gastrulation, by a movement suggestive of ingressions. These cell movements were observed not only at the posterior margin, where massive internalizations take place prior to the start of involution, but also in the center of the blastoderm, where internalizations of single cells prevail. Like the adjacent involuting mesendoderm, the posterior deep mesenchyme expresses anterior mesendoderm markers under the control of Nodal/activin signaling. Comparisons across vertebrates support the conclusion that endoderm is specified in two distinct temporal phases in the catshark as in all major osteichthyan lineages, in line with an ancient origin of a biphasic mode of endoderm specification in gnathostomes. They also highlight unexpected similarities with amniotes, such as the occurrence of cell ingressions from the superficial layer prior to gastrulation. These similarities may correspond to homoplastic traits fixed separately in amniotes and chondrichthyans and related to the increase in egg yolk mass.

The WTX/AMER1 gene family: evolution, signature and function.

BACKGROUND: WTX is a novel gene mutated in a proportion of Wilms' tumors and in patients suffering from sclerosing bone dysplasia. On the molecular level WTX has been shown to act as an antagonist of canonical Wnt/ß-catenin signaling in fish and mammals thus linking it to an essential pathway involved in normal development and cancer formation. Interestingly, WTX seems to also localize to an intranuclear component called paraspeckles. In spite of the growing interest of molecular biologists in WTX, little is known about its paralogs and its phylogenetic history. RESULTS: Using the amino-acid sequence of WTX/AMER1 as a tool for the assignment of orthology and paralogy, we here identify two novel proteins, AMER2 and AMER3, as "WTX" related. This Amer gene family is present in all currently available vertebrate genome sequences, but not invertebrate genomes and is characterized by six conserved blocks of sequences. The phylogenetic analysis suggests that the protoAmer gene originated early in the vertebrate lineage and was then duplicated due to whole genome duplications (WGD) giving rise to the three different Amer genes. CONCLUSION: Our study represents the first phylogenetic analysis of Amer genes and reveals a new vertebrate specific gene family that is likely to have played an important role in the evolution of this subphylum. Divergent and conserved molecular functions of Wtx/Amer1, Amer2 and Amer3 are discussed.

Expression patterns of the Wtx/Amer gene family during mouse embryonic development.

WTX/AMER1 is a novel negative regulator of the WNT/beta-catenin pathway with mutations detected in Wilms' tumors and an X-linked sclerosing bone dysplasia. WTX/AMER1 (Fam123b) shares several domains of homology with two other recently identified proteins: AMER2 (Fam123a) and AMER3 (Fam123c). Here, we describe an in-depth expression analysis of all three members of this gene family during mouse embryonic development. All genes were strongly expressed in the central as well as the peripheral nervous system, thus suggesting important roles of this gene family during neurogenesis. Specific expression was found in the retina, inner ear, and nasal epithelium. Outside of the nervous system Wtx/Amer1 showed the broadest expression domains including cephalic and limb mesenchyme, skeletal muscle, bladder, gonads, lung bud, salivary glands, and the kidneys. The widespread expression pattern of Wtx/Amer1, together with its role as a modulator of the Wnt signaling pathway, suggest that Wtx/Amer1 serves pleiotropic roles during mammalian organogenesis.

Reactivation of Snail genes in renal fibrosis and carcinomas: a process of reversed embryogenesis?

While the activity of Snail genes is required during embryonic development for the formation of different tissues and organs, they must be repressed in the adult in order to maintain epithelial integrity and homeostasis. Indeed, pathological activation of Snail in epithelial tumors induces malignancy and the recurrence of tumors. Here we show that in dedifferentiated areas of human renal carcinomas, Snail undergoes a process of reactivation. In addition to tumor progression, renal fibrosis is also linked to the activity of Snail genes and indeed, reactivation of Snail in the adult kidney is sufficient to induce fibrosis. Thus, Snail genes illustrate a paradigm whereby reactivation of crucial embryonic genes in adult tissues provokes the onset of devastating diseases.

Snail activation disrupts tissue homeostasis and induces fibrosis in the adult kidney.

During embryonic development, the kidney epithelium originates from cells that undergo a mesenchymal to epithelial transition (MET). The reverse process, epithelium to mesenchyme transition (EMT), has been implicated in epithelial tumor progression and in the fibrosis that leads to end-stage kidney failure. Snail transcription factors induce both natural and pathological EMT, but their implication in renal development and disease is still unclear. We show that Snail genes are downregulated during the MET that occurs during renal development and that this is correlated with Cadherin-16 expression. Snail suppresses Cadherin-16 via the direct repression of the kidney differentiation factor HNF-1beta, a novel route by which Snail disrupts epithelial homeostasis. Indeed, Snail activation is sufficient to induce EMT and kidney fibrosis in adult transgenic mice. Significantly, Snail is also activated in patients with renal fibrosis. Thus, Snail expression is suppressed during renal development and it must remain silent in the mature kidney where its aberrant activation leads to fibrosis.

Differential regulation of gelatinase A and B and TIMP-1 and -2 by TNFalpha and HIV virions in astrocytes.

Changes in the fine balance between matrix metalloproteinases and their tissue inhibitors, which drives extracellular matrix turnover, may be critical to central nervous system inflammation in HIV infection as well as in neurotoxicity. Although they do not produce virus when infected by HIV, astrocytes may be directly affected by the virion, because some viral proteins are known to transduce signaling in brain cells and are also sensitive to the major proinflammatory cytokine TNFalpha. We therefore studied the effects of HIV and TNFalpha on MMP-2, MMP-9 and their inhibitors, TIMP-1 and TIMP-2, in astrocytes, by zymography and ELISA, respectively, or by RT-PCR for both of them. HIV slightly increased the production of pro-MMP-2 and pro-MMP-9 by astrocytes, in a dose-dependent manner. TNFalpha strongly induced pro-MMP-9. TIMP-1 and TIMP-2 levels were affected only slightly, if at all, by HIV and TNFalpha. Thus, astrocyte/HIV contact may lead to extracellular matrix activation, which may be strongly amplified by the inflammatory response. Our data strongly suggest that, besides their physiological production of MMP-2, astrocytes would be a major source of MMP-9 in the inflamed brain.