Ancient deuterostome origins of vertebrate brain signalling centres

Neuroectodermal signalling centres induce and pattern many novel vertebrate brain structures but are absent, or divergent, in invertebrate chordates. This has led to the idea that signalling-centre genetic programs were first assembled in stem vertebrates and potentially drove morphological innovations of the brain. However, this scenario presumes that extant cephalochordates accurately represent ancestral chordate characters, which has not been tested using close chordate outgroups. Here we report that genetic programs homologous to three vertebrate signalling centres-the anterior neural ridge, zona limitans intrathalamica and isthmic organizer-are present in the hemichordate Saccoglossus kowalevskii. Fgf8/17/18 (a single gene homologous to vertebrate Fgf8, Fgf17 and Fgf18), sfrp1/5, hh and wnt1 are expressed in vertebrate-like arrangements in hemichordate ectoderm, and homologous genetic mechanisms regulate ectodermal patterning in both animals. We propose that these genetic programs were components of an unexpectedly complex, ancient genetic regulatory scaffold for deuterostome body patterning that degenerated in amphioxus and ascidians, but was retained to pattern divergent structures in hemichordates and vertebrates.     

The molecular nature of the zebrafish tail organizer

Based on grafting experiments, Mangold and Spemann showed the dorsal blastopore lip of an amphibian gastrula to be able to induce a secondary body axis. The equivalent of this organizer region has been identified in different vertebrates including teleosts. However, whereas the graft can induce ectopic head and trunk, endogenous and ectopic axes fuse in the posterior part of the body, raising the question of whether a distinct organizer region is necessary for tail development. Here we reveal, by isochronic and heterochronic transplantation, the existence of a tail organizer deriving from the ventral margin of the zebrafish embryo, which is independent of the dorsal Spemann organizer. Loss-of-function experiments reveal that bone morphogenetic protein (BMP), Nodal and Wnt8 signalling pathways are required for tail development. Moreover, stimulation of naive cells by a combination of BMP, Nodal and Wnt8 mimics the tail-organizing activity of the ventral margin and induces surrounding tissues to become tail. In contrast to induction of the vertebrate head, known to result from the triple inhibition of BMP, Nodal and Wnt, here we show that induction of the tail results from the triple stimulation of BMP, Nodal and Wnt8 signalling pathways.     

Conservation and elaboration of Hox gene regulation during evolution of the vertebrate head

The comparison of Hox genes between vertebrates and their closest invertebrate relatives (amphioxus and ascidia) highlights two derived features of Hox genes in vertebrates: duplication of the Hox gene cluster, and an elaboration of Hox expression patterns and roles compared with non-vertebrate chordates. We have investigated how new expression domains and their associated developmental functions evolved, by testing the cis-regulatory activity of genomic DNA fragments from the cephalochordate amphioxus Hox cluster in transgenic mouse and chick embryos. Here we present evidence for the conservation of cis-regulatory mechanisms controlling gene expression in the neural tube for half a billion years of evolution, including a dependence on retinoic acid signalling. We also identify amphioxus Hox gene regulatory elements that drive spatially localized expression in vertebrate neural crest cells, in derivatives of neurogenic placodes and in branchial arches, despite the fact that cephalochordates lack both neural crest and neurogenic placodes. This implies an elaboration of cis-regulatory elements in the Hox gene cluster of vertebrate ancestors during the evolution of craniofacial patterning.     

Repressor activity of Headless/Tcf3 is essential for vertebrate head formation

The vertebrate organizer can induce a complete body axis when transplanted to the ventral side of a host embryo by virtue of its distinct head and trunk inducing properties. Wingless/Wnt antagonists secreted by the organizer have been identified as head inducers. Their ectopic expression can promote head formation, whereas ectopic activation of Wnt signalling during early gastrulation blocks head formation. These observations suggest that the ability of head inducers to inhibit Wnt signalling during formation of anterior structures is what distinguishes them from trunk inducers that permit the operation of posteriorizing Wnt signals. Here we describe the zebrafish headless (hdl) mutant and show that its severe head defects are due to a mutation in T-cell factor-3 (Tcf3), a member of the Tcf/Lef family. Loss of Tcf3 function in the hdl mutant reveals that hdl represses Wnt target genes. We provide genetic evidence that a component of the Wnt signalling pathway is essential in vertebrate head formation and patterning.     

MicroRNA control of Nodal signalling

MicroRNAs are crucial modulators of gene expression, yet their involvement as effectors of growth factor signalling is largely unknown. Ligands of the transforming growth factor-beta superfamily are essential for development and adult tissue homeostasis. In early Xenopus embryos, signalling by the transforming growth factor-beta ligand Nodal is crucial for the dorsal induction of the Spemann's organizer. Here we report that Xenopus laevis microRNAs miR-15 and miR-16 restrict the size of the organizer by targeting the Nodal type II receptor Acvr2a. Endogenous miR-15 and miR-16 are ventrally enriched as they are negatively regulated by the dorsal Wnt/beta-catenin pathway. These findings exemplify the relevance of microRNAs as regulators of early embryonic patterning acting at the crossroads of fundamental signalling cascades.     

An LDL-receptor-related protein mediates Wnt signalling in mice

Wnt genes comprise a large family of secreted polypeptides that are expressed in spatially and tissue-restricted patterns during vertebrate embryonic development. Mutational analysis in mice has shown the importance of Wnts in controlling diverse developmental processes such as patterning of the body axis, central nervous system and limbs, and the regulation of inductive events during organogenesis. Although many components of the Wnt signalling pathway have been identified, little is known about how Wnts and their cognate Frizzled receptors signal to downstream effector molecules. Here we present evidence that a new member of the low-density lipoprotein (LDL)-receptor-related protein family, LRP6 (ref. 3), is critical for Wnt signalling in mice. Embryos homozygous for an insertion mutation in the LRP6 gene exhibit developmental defects that are a striking composite of those caused by mutations in individual Wnt genes. Furthermore, we show a genetic enhancement of a Wnt mutant phenotype in mice lacking one functional copy of LRP6. Together, our results support a broad role for LRP6 in the transduction of several Wnt signals in mammals.     

Dependence of Drosophila wing imaginal disc cytonemes on Decapentaplegic

The anterior/posterior (A/P) and dorsal/ventral (D/V) compartment borders that subdivide the wing imaginal discs of Drosophila third instar larvae are each associated with a developmental organizer. Decapentaplegic (Dpp), a member of the transforming growth factor-beta (TGF-beta) superfamily, embodies the activity of the A/P organizer. It is produced at the A/P organizer and distributes in a gradient of decreasing concentration to regulate target genes, functioning non-autonomously to regulate growth and patterning of both the anterior and posterior compartments. Wingless (Wg) is produced at the D/V organizer and embodies its activity. The mechanisms that distribute Dpp and Wg are not known, but proposed mechanisms include extracellular diffusion, successive transfers between neighbouring cells, vesicle-mediated movement, and direct transfer via cytonemes. Cytonemes are actin-based filopodial extensions that have been found to orient towards the A/P organizer from outlying cells. Here we show that in the wing disc, cytonemes orient towards both the A/P and D/V organizers, and that their presence and orientation correlates with Dpp signalling. We also show that the Dpp receptor, Thickveins (Tkv), is present in punctae that move along cytonemes. These observations are consistent with a role for cytonemes in signal transduction.     

Protochordate amphioxus is an emerging model organism for comparative immunology

Protochordate amphioxus is an extant invertebrate regarded quite recently as a basal chordate. It has a vertebrate-like body plan including a circulation system with an organization similar to that of vertebrates. However, amphioxus is less complex than vertebrates for having a genome uncomplicated by extensive genomic duplication, and lacking lymphoid organs and free circulating blood cells. Recent studies on immunity have demonstrated the presence in amphioxus of both the constituent elements of key molecules involved in adaptive immunity such as proto-major histocompatibility complex (proto-MHC), V region-containing chitin-binding protein (VCBP) and V and C domain-bearing protein (VCP), and the complement system operating via the alternative and lectin pathways resembling those seen in vertebrates. In addition, the acute phase response profile in amphioxus has been shown to be similar to that observed in vertebrates. These findings together with the relative structural and genomic simplicity make amphioxus an ideal organism for gaining insights into the origin and evolution of the vertebrate immune system, especially adaptive immunity, and the composition and mechanisms of the vertebrate innate immunity.     

Wnt-11 activation of a non-canonical Wnt signalling pathway is required for cardiogenesis

Formation of the vertebrate heart requires a complex interplay of several temporally regulated signalling cascades. In Xenopus laevis, cardiac specification occurs during gastrulation and requires signals from the dorsal lip and underlying endoderm. Among known Xenopus Wnt genes, only Wnt-11 shows a spatiotemporal pattern of expression that correlates with cardiac specification, which indicates that Wnt-11 may be involved in heart development. Here we show, through loss- and gain-of-function experiments, that XWnt-11 is required for heart formation in Xenopus embryos and is sufficient to induce a contractile phenotype in embryonic explants. Treating the mouse embryonic carcinoma stem cell line P19 with murine Wnt-11 conditioned medium triggers cardiogenesis, which indicates that the function of Wnt-11 in heart development has been conserved in higher vertebrates. XWnt-11 mediates this effect by non-canonical Wnt signalling, which is independent of beta-catenin and involves protein kinase C and Jun amino-terminal kinase. Our results indicate that the cardiac developmental program requires non-canonical Wnt signal transduction.     

Distalization of the Drosophila leg by graded EGF-receptor activity

Arthropods and higher vertebrates both possess appendages, but these are morphologically distinct and the molecular mechanisms regulating patterning along their proximodistal axis (base to tip) are thought to be quite different. In Drosophila, gene expression along this axis is thought to be controlled primarily by a combination of transforming growth factor-beta (TGF-beta) and Wnt signalling from sources of ligands, Decapentaplegic (Dpp) and Wingless (Wg), in dorsal and ventral stripes, respectively. In vertebrates, however, proximodistal patterning is regulated by receptor tyrosine kinase (RTK) activity from a source of ligands, fibroblast growth factors (FGFs), at the tip of the limb bud. Here I revise our understanding of limb development in flies and show that the distal region is actually patterned by a distal-to-proximal gradient of RTK activity, established by a source of epidermal growth factor (EGF)-related ligands at the presumptive tip. This similarity between proximodistal patterning in vertebrates and flies supports previous suggestions of an evolutionary relationship between appendages/body-wall outgrowths in animals.     

Maternal nodal and zebrafish embryogenesis

In fish and amphibians, the dorsal axis is specified by the asymmetric localization of maternally provided components of the Wnt signalling pathway. Gore et al. suggest that the Nodal signal Squint (Sqt) is required as a maternally provided dorsal determinant in zebrafish. Here we test their proposal and show that the maternal activities of sqt and the related Nodal gene cyclops (cyc) are not required for dorsoventral patterning.     

人类抗体重排机制的起源

 重组激活基因（RAG）蛋白介导的抗体重排是脊椎动物适应性免疫系统的核心,抗体重排机制的起源一直是免疫学研究的热点。本研究以有活化石之称的文昌鱼为对象,对多个文昌鱼基因组草图进行深度信息学分析,发现了一个全新DNA转座子ProtoRAG。进一步功能研究表明,ProtoRAG编码的蛋白能够介导自身的转座和宿主DNA的重排,其作用机制与人类抗体蛋白介导的抗体重排机制基本一致。因此,ProtoRAG就是研究人员长期搜寻的祖先RAG转座子的“分子活化石”,该发现为诺贝尔生理学或医学奖获得者利根川进提出的“抗体重排机制转座子起源”假说提供了最有力和最直接的证据。     

Unexpected complexity of the Wnt gene family in a sea anemone

The Wnt gene family encodes secreted signalling molecules that control cell fate in animal development and human diseases. Despite its significance, the evolution of this metazoan-specific protein family is unclear. In vertebrates, twelve Wnt subfamilies were defined, of which only six have counterparts in Ecdysozoa (for example, Drosophila and Caenorhabditis). Here, we report the isolation of twelve Wnt genes from the sea anemone Nematostella vectensis, a species representing the basal group within cnidarians. Cnidarians are diploblastic animals and the sister-group to bilaterian metazoans. Phylogenetic analyses of N. vectensis Wnt genes reveal a thus far unpredicted ancestral diversity within the Wnt family. Cnidarians and bilaterians have at least eleven of the twelve known Wnt gene subfamilies in common; five subfamilies appear to be lost in the protostome lineage. Expression patterns of Wnt genes during N. vectensis embryogenesis indicate distinct roles of Wnts in gastrulation, resulting in serial overlapping expression domains along the primary axis of the planula larva. This unexpectedly complex inventory of Wnt family signalling factors evolved in early multi-cellular animals about 650 million years (Myr) ago, predating the Cambrian explosion by at least 100 Myr (refs 5, 8). It emphasizes the crucial function of Wnt genes in the diversification of eumetazoan body plans.     

Planar cell polarity signalling controls cell division orientation during zebrafish gastrulation

Oriented cell division is an integral part of pattern development in processes ranging from asymmetric segregation of cell-fate determinants to the shaping of tissues. Despite proposals that it has an important function in tissue elongation, the mechanisms regulating division orientation have been little studied outside of the invertebrates Caenorhabditis elegans and Drosophila melanogaster. Here, we have analysed mitotic divisions during zebrafish gastrulation using in vivo confocal imaging and found that cells in dorsal tissues preferentially divide along the animal-vegetal axis of the embryo. Establishment of this animal-vegetal polarity requires the Wnt pathway components Silberblick/Wnt11, Dishevelled and Strabismus. Our findings demonstrate an important role for non-canonical Wnt signalling in oriented cell division during zebrafish gastrulation, and indicate that oriented cell division is a driving force for axis elongation. Furthermore, we propose that non-canonical Wnt signalling has a conserved role in vertebrate axis elongation, orienting both cell intercalation and mitotic division.     

青岛文昌鱼hedgehog及两个hedgehog-like片段的克隆

采用RT-PCR方法获得了青岛文昌鱼2497bp长的hedgehog基因片段,其所编码的氨基酸序列与多种生物hh基因家族成员的相应片段显示了较高的同源性.同时获得2个含有部分hedgehog基因片段的序列hedgehog-like-1和hedgehog-like-2,提示文昌鱼中发生hedgehog基因倍增过程和有多拷贝hh基因存在的可能性.为研究hh基因的分子进化提供了线索.