Molecular Characterization of the Gastrula in the Turtle Emys orbicularis: An Evolutionary Perspective on Gastrulation

Due to the presence of a blastopore as in amphibians, the turtle has been suggested to exemplify a transition form from an amphibian- to an avian-type gastrulation pattern. In order to test this hypothesis and gain insight into the emergence of the unique characteristics of amniotes during gastrulation, we have performed the first molecular characterization of the gastrula in a reptile, the turtle Emys orbicularis. The study of Brachyury, Lim1, Otx2 and Otx5 expression patterns points to a highly conserved dynamic of expression with amniote model organisms and makes it possible to identify the site of mesoderm internalization, which is a long-standing issue in reptiles. Analysis of Brachyury expression also highlights the presence of two distinct phases, less easily recognizable in model organisms and respectively characterized by an early ring-shaped and a later bilateral symmetrical territory. Systematic comparisons with tetrapod model organisms lead to new insights into the relationships of the blastopore/blastoporal plate system shared by all reptiles, with the blastopore of amphibians and the primitive streak of birds and mammals. The biphasic Brachyury expression pattern is also consistent with recent models of emergence of bilateral symmetry, which raises the question of its evolutionary significance

Analyses of zebrafish and Xenopus oocyte maturation reveal conserved and diverged features of translational regulation of maternal cyclin B1 mRNA

<p>Abstract</p> <p>Background</p> <p>Vertebrate development relies on the regulated translation of stored maternal mRNAs, but how these regulatory mechanisms may have evolved to control translational efficiency of individual mRNAs is poorly understood. We compared the translational regulation and polyadenylation of the cyclin B1 mRNA during zebrafish and <it>Xenopus </it>oocyte maturation. Polyadenylation and translational activation of cyclin B1 mRNA is well characterized during <it>Xenopus </it>oocyte maturation. Specifically, <it>Xenopus </it>cyclin B1 mRNA is polyadenylated and translationally activated during oocyte maturation by proteins that recognize the conserved AAUAAA hexanucleotide and U-rich Cytoplasmic Polyadenylation Elements (CPEs) within cyclin B1 mRNA's 3'<b>U</b>n<b>T</b>ranslated <b>R</b>egion (3'<b>UTR</b>).</p> <p>Results</p> <p>The zebrafish cyclin B1 mRNA was polyadenylated during zebrafish oocyte maturation. Furthermore, the zebrafish cyclin B1 mRNA's 3'UTR was sufficient to stimulate translation of a reporter mRNA during zebrafish oocyte maturation. This stimulation required both AAUAAA and U-rich CPE-like sequences. However, in contrast to AAUAAA, the positions and sequences of the functionally defined CPEs were poorly conserved between <it>Xenopus </it>and zebrafish cyclin B1 mRNA 3'UTRs. To determine whether these differences were relevant to translation efficiency, we analyzed the translational activity of reporter mRNAs containing either the zebrafish or <it>Xenopus </it>cyclin B1 mRNA 3'UTRs during both zebrafish and <it>Xenopus </it>oocyte maturation. The zebrafish cyclin B1 3'UTR was quantitatively less effective at stimulating polyadenylation and translation compared to the <it>Xenopus </it>cyclin B1 3'UTR during both zebrafish and <it>Xenopus </it>oocyte maturation.</p> <p>Conclusion</p> <p>Although the factors that regulate translation of maternal mRNAs are highly conserved, the target sequences and overall sequence architecture within the 3'UTR of the cyclin B1 mRNA have diverged to affect translational efficiency, perhaps to optimize levels of cyclin B1 protein required by these different species during their earliest embryonic cell divisions.</p

The TATA-binding protein regulates maternal mRNA degradation and differential zygotic transcription in zebrafish

Early steps of embryo development are directed by maternal gene products and trace levels of zygotic gene activity in vertebrates. A major activation of zygotic transcription occurs together with degradation of maternal mRNAs during the midblastula transition in several vertebrate systems. How these processes are regulated in preparation for the onset of differentiation in the vertebrate embryo is mostly unknown. Here, we studied the function of TATA-binding protein (TBP) by knock down and DNA microarray analysis of gene expression in early embryo development. We show that a subset of polymerase II-transcribed genes with ontogenic stage-dependent regulation requires TBP for their zygotic activation. TBP is also required for limiting the activation of genes during development. We reveal that TBP plays an important role in the degradation of a specific subset of maternal mRNAs during late blastulation/early gastrulation, which involves targets of the miR-430 pathway. Hence, TBP acts as a specific regulator of the key processes underlying the transition from maternal to zygotic regulation of embryogenesis. These results implicate core promoter recognition as an additional level of differential gene regulation during development

Congenital Hydrocephalus and Abnormal Subcommissural Organ Development in Sox3 Transgenic Mice

Congenital hydrocephalus (CH) is a life-threatening medical condition in which excessive accumulation of CSF leads to ventricular expansion and increased intracranial pressure. Stenosis (blockage) of the Sylvian aqueduct (Aq; the narrow passageway that connects the third and fourth ventricles) is a common form of CH in humans, although the genetic basis of this condition is unknown. Mouse models of CH indicate that Aq stenosis is associated with abnormal development of the subcommmissural organ (SCO) a small secretory organ located at the dorsal midline of the caudal diencephalon. Glycoproteins secreted by the SCO generate Reissner's fibre (RF), a thread-like structure that descends into the Aq and is thought to maintain its patency. However, despite the importance of SCO function in CSF homeostasis, the genetic program that controls SCO development is poorly understood. Here, we show that the X-linked transcription factor SOX3 is expressed in the murine SCO throughout its development and in the mature organ. Importantly, overexpression of Sox3 in the dorsal diencephalic midline of transgenic mice induces CH via a dose-dependent mechanism. Histological, gene expression and cellular proliferation studies indicate that Sox3 overexpression disrupts the development of the SCO primordium through inhibition of diencephalic roof plate identity without inducing programmed cell death. This study provides further evidence that SCO function is essential for the prevention of hydrocephalus and indicates that overexpression of Sox3 in the dorsal midline alters progenitor cell differentiation in a dose-dependent manner

Spatial Analysis of Expression Patterns Predicts Genetic Interactions at the Mid-Hindbrain Boundary

The isthmic organizer mediating differentiation of mid- and hindbrain during vertebrate development is characterized by a well-defined pattern of locally restricted gene expression domains around the mid-hindbrain boundary (MHB). This pattern is established and maintained by a regulatory network between several transcription and secreted factors that is not yet understood in full detail. In this contribution we show that a Boolean analysis of the characteristic spatial gene expression patterns at the murine MHB reveals key regulatory interactions in this network. Our analysis employs techniques from computational logic for the minimization of Boolean functions. This approach allows us to predict also the interplay of the various regulatory interactions. In particular, we predict a maintaining, rather than inducing, effect of Fgf8 on Wnt1 expression, an issue that remained unclear from published data. Using mouse anterior neural plate/tube explant cultures, we provide experimental evidence that Fgf8 in fact only maintains but does not induce ectopic Wnt1 expression in these explants. In combination with previously validated interactions, this finding allows for the construction of a regulatory network between key transcription and secreted factors at the MHB. Analyses of Boolean, differential equation and reaction-diffusion models of this network confirm that it is indeed able to explain the stable maintenance of the MHB as well as time-courses of expression patterns both under wild-type and various knock-out conditions. In conclusion, we demonstrate that similar to temporal also spatial expression patterns can be used to gain information about the structure of regulatory networks. We show, in particular, that the spatial gene expression patterns around the MHB help us to understand the maintenance of this boundary on a systems level

Coe Genes Are Expressed in Differentiating Neurons in the Central Nervous System of Protostomes

Genes of the coe (collier/olfactory/early B-cell factor) family encode Helix-Loop-Helix transcription factors that are widely conserved in metazoans and involved in many developmental processes, neurogenesis in particular. Whereas their functions during vertebrate neural tube formation have been well documented, very little is known about their expression and role during central nervous system (CNS) development in protostomes. Here we characterized the CNS expression of coe genes in the insect Drosophila melanogaster and the polychaete annelid Platynereis dumerilii, which belong to different subgroups of protostomes and show strikingly different modes of development. In the Drosophila ventral nerve cord, we found that the Collier-expressing cells form a subpopulation of interneurons with diverse molecular identities and neurotransmitter phenotypes. We also demonstrate that collier is required for the proper differentiation of some interneurons belonging to the Eve-Lateral cluster. In Platynereis dumerilii, we cloned a single coe gene, Pdu-coe, and found that it is exclusively expressed in post mitotic neural cells. Using an original technique of in silico 3D registration, we show that Pdu-coe is co-expressed with many different neuronal markers and therefore that, like in Drosophila, its expression defines a heterogeneous population of neurons with diverse molecular identities. Our detailed characterization and comparison of coe gene expression in the CNS of two distantly-related protostomes suggest conserved roles of coe genes in neuronal differentiation in this clade. As similar roles have also been observed in vertebrates, this function was probably already established in the last common ancestor of all bilaterians

Adult zebrafish as a model organism for behavioural genetics

Recent research has demonstrated the suitability of adult zebrafish to model some aspects of complex behaviour. Studies of reward behaviour, learning and memory, aggression, anxiety and sleep strongly suggest that conserved regulatory processes underlie behaviour in zebrafish and mammals. The isolation and molecular analysis of zebrafish behavioural mutants is now starting, allowing the identification of novel behavioural control genes. As a result of this, studies of adult zebrafish are now helping to uncover the genetic pathways and neural circuits that control vertebrate behaviour