Evolution of the new head by gradual acquisition of neural crest regulatory circuits.

The neural crest, an embryonic stem-cell population, is a vertebrate innovation that has been proposed to be a key component of the 'new head', which imbued vertebrates with predatory behaviour1,2. Here, to investigate how the evolution of neural crest cells affected the vertebrate body plan, we examined the molecular circuits that control neural crest development along the anteroposterior axis of a jawless vertebrate, the sea lamprey. Gene expression analysis showed that the cranial subpopulation of the neural crest of the lamprey lacks most components of a transcriptional circuit that is specific to the cranial neural crest in amniotes and confers the ability to form craniofacial cartilage onto non-cranial neural crest subpopulations3. Consistent with this, hierarchical clustering analysis revealed that the transcriptional profile of the lamprey cranial neural crest is more similar to the trunk neural crest of amniotes. Notably, analysis of the cranial neural crest in little skate and zebrafish embryos demonstrated that the transcriptional circuit that is specific to the cranial neural crest emerged via the gradual addition of network components to the neural crest of gnathostomes, which subsequently became restricted to the cephalic region. Our results indicate that the ancestral neural crest at the base of the vertebrate lineage possessed a trunk-like identity. We propose that the emergence of the cranial neural crest, by progressive assembly of an axial-specific regulatory circuit, allowed the elaboration of the new head during vertebrate evolution.

Inner ear development in cyclostomes and evolution of the vertebrate semicircular canals.

Jawed vertebrates have inner ears with three semicircular canals, the presence of which has been used as a key to understanding evolutionary relationships. Ostracoderms, the jawless stem gnathostomes, had only two canals and lacked the lateral canal1-3. Lampreys, which are modern cyclostomes, are generally thought to possess two semicircular canals whereas the hagfishes-which are also cyclostomes-have only a single canal, which used to be regarded as a more primitive trait1,4. However, recent molecular and developmental analyses have strongly supported the monophyly of cyclostomes5-7, which has left the evolutionary trajectory of the vertebrate inner ear unclear8. Here we show the differentiation of the otic vesicle of the lamprey Lethenteron camtschaticum and inshore hagfish Eptatretus burgeri. This is the first time, to our knowledge, that the development of the hagfish inner ear is reported. We found that canal development in the lamprey starts with two depressions-which is reminiscent of the early developmental pattern of the inner ear in modern gnathostomes. These cyclostome otic vesicles show a pattern of expression of regulatory genes, including OTX genes, that is comparable to that of gnathosomes. Although two depressions appear in the lamprey vesicle, they subsequently fuse to form a single canal that is similar to that of hagfishes. Complete separation of the depressions results in anterior and posterior canals in gnathostomes. The single depression of the vesicle in hagfishes thus appears to be a secondarily derived trait. Furthermore, the lateral canal in crown gnathostomes was acquired secondarily-not by de novo acquisition of an OTX expression domain, but by the evolution of a developmental program downstream of the OTX genes.

A Notch-regulated proliferative stem cell zone in the developing spinal cord is an ancestral vertebrate trait.

Vertebrates have evolved the most sophisticated nervous systems we know. These differ from the nervous systems of invertebrates in several ways, including the evolution of new cell types, and the emergence and elaboration of patterning mechanisms to organise cells in time and space. Vertebrates also generally have many more cells in their central nervous systems than invertebrates, and an increase in neural cell number may have contributed to the sophisticated anatomy of the brain and spinal cord. Here, we study how increased cell number evolved in the vertebrate central nervous system, investigating the regulation of cell proliferation in the lamprey spinal cord. Markers of proliferation show that a ventricular progenitor zone is found throughout the lamprey spinal cord. We show that inhibition of Notch signalling disrupts the maintenance of this zone. When Notch is blocked, progenitor cells differentiate precociously, the proliferative ventricular zone is lost and differentiation markers become expressed throughout the spinal cord. Comparison with other chordates suggests that the emergence of a persistent Notch-regulated proliferative progenitor zone was a crucial step for the evolution of vertebrate spinal cord complexity.

Evidence from cyclostomes for complex regionalization of the ancestral vertebrate brain.

The vertebrate brain is highly complex, but its evolutionary origin remains elusive. Because of the absence of certain developmental domains generally marked by the expression of regulatory genes, the embryonic brain of the lamprey, a jawless vertebrate, had been regarded as representing a less complex, ancestral state of the vertebrate brain. Specifically, the absence of a Hedgehog- and Nkx2.1-positive domain in the lamprey subpallium was thought to be similar to mouse mutants in which the suppression of Nkx2-1 leads to a loss of the medial ganglionic eminence. Here we show that the brain of the inshore hagfish (Eptatretus burgeri), another cyclostome group, develops domains equivalent to the medial ganglionic eminence and rhombic lip, resembling the gnathostome brain. Moreover, further investigation of lamprey larvae revealed that these domains are also present, ruling out the possibility of convergent evolution between hagfish and gnathostomes. Thus, brain regionalization as seen in crown gnathostomes is not an evolutionary innovation of this group, but dates back to the latest vertebrate ancestor before the divergence of cyclostomes and gnathostomes more than 500 million years ago.

Development of the lamprey velum and implications for the evolution of the vertebrate jaw.

BACKGROUND: The vertebrate jaw is thought to have evolved through developmental modification of the mandibular arch. An extant jawless vertebrate, the lamprey, possesses a structure called "velum"-a mandibular arch derivative-in addition to the oral apparatus. This leads us to assess the velum's possible contribution to the evolution of jaws. RESULTS: The velar muscles develop from progenitor cells distinct from those from which the oral muscles develop. In addition, the oral and velar regions originate from the different sub-population of the trigeminal neural crest cells (NCCs): the former region receives NCCs from the midbrain, whereas the latter region receives NCCs from the anterior hindbrain. The expression of patterning genes (eg, DlxA and MsxA) is activated at a later developmental stage in the velum compared to the oral region, and more importantly, in different cells from those in the oral region. CONCLUSION: The lamprey mandibular arch consists of two developmental units: the anterior oral unit and the posterior velar unit. Because structural elements of the lamprey velum may be homologous to the jaw, the evolution of vertebrate jaws may have occurred by the velum being released from its functional roles in feeding or respiration in jawless vertebrates.

Evolution of Snail-mediated regulation of neural crest and placodes from an ancient role in bilaterian neurogenesis.

A major challenge in vertebrate evolution is to identify the gene regulatory mechanisms that facilitated the origin of neural crest cells and placodes from ancestral precursors in invertebrates. Here, we show in lamprey, a primitively jawless vertebrate, that the transcription factor Snail is expressed simultaneously throughout the neural plate, neural plate border, and pre-placodal ectoderm in the early embryo and is then upregulated in the CNS throughout neurogenesis. Using CRISPR/Cas9-mediated genome editing, we demonstrate that Snail plays functional roles in all of these embryonic domains or their derivatives. We first show that Snail patterns the neural plate border by repressing lateral expansion of Pax3/7 and activating nMyc and ZicA. We also present evidence that Snail is essential for DlxB-mediated establishment of the pre-placodal ectoderm but is not required for SoxB1a expression during formation of the neural plate proper. At later stages, Snail regulates formation of neural crest-derived and placode-derived PNS neurons and controls CNS neural differentiation in part by promoting cell survival. Taken together with established functions of invertebrate Snail genes, we identify a pan-bilaterian mechanism that extends to jawless vertebrates for regulating neurogenesis that is dependent on Snail transcription factors. We propose that ancestral vertebrates deployed an evolutionarily conserved Snail expression domain in the CNS and PNS for neurogenesis and then acquired derived functions in neural crest and placode development by recruitment of regulatory genes downstream of neuroectodermal Snail activity. Our results suggest that Snail regulatory mechanisms in vertebrate novelties such as the neural crest and placodes may have emerged from neurogenic roles that originated early in bilaterian evolution.

A Hox regulatory network of hindbrain segmentation is conserved to the base of vertebrates.

A defining feature governing head patterning of jawed vertebrates is a highly conserved gene regulatory network that integrates hindbrain segmentation with segmentally restricted domains of Hox gene expression. Although non-vertebrate chordates display nested domains of axial Hox expression, they lack hindbrain segmentation. The sea lamprey, a jawless fish, can provide unique insights into vertebrate origins owing to its phylogenetic position at the base of the vertebrate tree. It has been suggested that lamprey may represent an intermediate state where nested Hox expression has not been coupled to the process of hindbrain segmentation. However, little is known about the regulatory network underlying Hox expression in lamprey or its relationship to hindbrain segmentation. Here, using a novel tool that allows cross-species comparisons of regulatory elements between jawed and jawless vertebrates, we report deep conservation of both upstream regulators and segmental activity of enhancer elements across these distant species. Regulatory regions from diverse gnathostomes drive segmental reporter expression in the lamprey hindbrain and require the same transcriptional inputs (for example, Kreisler (also known as Mafba), Krox20 (also known as Egr2a)) in both lamprey and zebrafish. We find that lamprey hox genes display dynamic segmentally restricted domains of expression; we also isolated a conserved exonic hox2 enhancer from lamprey that drives segmental expression in rhombomeres 2 and 4. Our results show that coupling of Hox gene expression to segmentation of the hindbrain is an ancient trait with origin at the base of vertebrates that probably led to the formation of rhombomeric compartments with an underlying Hox code.

Gliogenesis in lampreys shares gene regulatory interactions with oligodendrocyte development in jawed vertebrates.

Glial cells in the nervous system regulate and support many functions related to neuronal activity. Understanding how the vertebrate nervous system has evolved demands a greater understanding of the mechanisms controlling evolution and development of glial cells in basal vertebrates. Among vertebrate glia, oligodendrocytes form an insulating myelin layer surrounding axons of the central nervous system (CNS) in jawed vertebrates. Jawless vertebrates lack myelinated axons but it is unclear when oligodendrocytes or the regulatory mechanisms controlling their development evolved. To begin to investigate the evolution of mechanisms controlling glial development, we identified key genes required for the differentiation of oligodendrocytes in gnathostomes, including Nkx2.2, SoxE genes, and PDGFR, analyzed their expression, and used CRISPR/Cas9 genome editing to perturb their functions in a primitively jawless vertebrate, the sea lamprey. We show in lamprey that orthologs required for oligodendrocyte development in jawed vertebrates are expressed in the lamprey ventral neural tube, in similar locations where gnathostome oligodendrocyte precursor cells (OPC) originate. In addition, they appear to be under the control of conserved mechanisms that regulate OPC development in jawed vertebrates and may also function in gliogenesis. Our results suggest that although oligodendrocytes first emerged in jawed vertebrates, regulatory mechanisms required for their development predate the divergence of jawless and jawed vertebrates.

Species-specific contribution of volumetric growth and tissue convergence to posterior body elongation in vertebrates.

Posterior body elongation is a widespread mechanism propelling the generation of the metazoan body plan. The posterior growth model predicts that a posterior growth zone generates sufficient tissue volume to elongate the posterior body. However, there are energy supply-related differences between vertebrates in the degree to which growth occurs concomitantly with embryogenesis. By applying a multi-scalar morphometric analysis in zebrafish embryos, we show that posterior body elongation is generated by an influx of cells from lateral regions, by convergence-extension of cells as they exit the tailbud, and finally by a late volumetric growth in the spinal cord and notochord. Importantly, the unsegmented region does not generate additional tissue volume. Fibroblast growth factor inhibition blocks tissue convergence rather than volumetric growth, showing that a conserved molecular mechanism can control convergent morphogenesis through different cell behaviours. Finally, via a comparative morphometric analysis in lamprey, dogfish, zebrafish and mouse, we propose that elongation via posterior volumetric growth is linked to increased energy supply and is associated with an overall increase in volumetric growth and elongation.

Craniofacial development of hagfishes and the evolution of vertebrates.

Cyclostomes, the living jawless vertebrates including hagfishes and lampreys, represent the most basal lineage of vertebrates. Although the monophyly of cyclostomes has been supported by recent molecular analyses, the phenotypic traits of hagfishes, especially the lack of some vertebrate-defining features and the reported endodermal origin of the adenohypophysis, have been interpreted as hagfishes exhibiting a more ancestral state than those of all other vertebrates. Furthermore, the adult anatomy of hagfishes cannot be compared easily with that of lampreys. Here we describe the craniofacial development of a series of staged hagfish embryos, which shows that their adenohypophysis arises ectodermally, consistent with the molecular phylogenetic data. This finding also allowed us to identify a pan-cyclostome pattern, one not shared by jawed vertebrates. Comparative analyses indicated that many of the hagfish-specific traits can be explained by changes secondarily introduced into the hagfish lineage. We also propose a possibility that the pan-cyclostome pattern may reflect the ancestral programme for the craniofacial development of all living vertebrates.

The neural crest and origin of the neurocranium in vertebrates.

Cranium of jawed vertebrates is composed of dorsal moiety that encapsulates the brain, or the neurocranium, and the is called the neurocranium, and the ventral moiety, the viscerocranium, that supports the pharynx. In modern jawed vertebrates (crown gnathostomes), the viscerocranium is predominantly of neural crest origin, and for the neurocranium, the rostral part is derived from neural crest cells, whereas the posterior part from the mesoderm. In the cyclostome cranium, the mesoderm/neural crest boundary of the neurocranium used to be enigmatic, let alone the morphological comparison of neurocranial between two cyclostome groups, lampreys and hagfishes. By examining the hagfish development it has become clear that cyclostomes share a common craniofacial embryonic pattern that is not shared by modern gnathostomes, and cyclostome cranium can be compared among the group as developmental modular units with comparable mesoderm/neural crest boundary within the neuroranium. Also, the dual origin of the jawed vertebrate neurocranium has now turned out to represent a derived condition, and ancestrally, the neurocranium would likely have been predominantly of mesodermal origin. Enlargement of the forebrain and reorganization of the oral apparatus seem to have led to the involvement of the neural crest in the rostral neurocranium.

Lamprey neural crest migration is Snail-dependent and occurs without a differential shift in cadherin expression.

The acquisition of neural crest cells was a key step in the origin of the vertebrate body plan. An outstanding question is how neural crest cells acquired their ability to undergo an epithelial-mesenchymal transition (EMT) and migrate extensively throughout the vertebrate embryo. We tested if differential regulation of classical cadherins-a highly conserved feature of neural crest EMT and migration in jawed vertebrates-mediates these cellular behaviors in lamprey, a basal jawless vertebrate. Lamprey has single copies of the type I and type II classical cadherins (CadIA and CadIIA). CadIIA is expressed in premigratory neural crest, and requires the transcription factor Snail for proper expression, yet CadIA is never expressed in the neural tube during neural crest development, suggesting that differential regulation of classical cadherin expression is not required to initiate neural crest migration in basal vertebrates. We hypothesize that neural crest cells evolved by retention of regulatory programs linking distinct mesenchymal and multipotency properties, and emigrated from the neural tube without differentially regulating type I/type II cadherins. Our results point to the coupling of mesenchymal state and multipotency as a key event facilitating the origin of migratory neural crest cells.

Functional constraints on SoxE proteins in neural crest development: The importance of differential expression for evolution of protein activity.

Vertebrate SoxE genes (Sox8, 9, and 10) are key regulators of neural crest cell (NCC) development. These genes arose by duplication from a single SoxE gene in the vertebrate ancestor. Although SoxE paralogs are coexpressed early in NCC development, later, Sox9 is restricted to skeletogenic lineages in the head, and Sox10 to non-skeletogenic NCC in the trunk and head. When this subfunctionalization evolved and its possible role in the evolution of the neural crest are unknown. Sea lampreys are basal vertebrates that also possess three SoxE genes, while only a single SoxE is present in the cephalochordate amphioxus. In order to address the functional divergence of SoxE genes, and to determine if differences in their biochemical functions may be linked to changes in neural crest developmental potential, we examined the ability of lamprey and amphioxus SoxE genes to regulate differentiation of NCC derivatives in zebrafish colourless (cls) mutants lacking expression of sox10. Our findings suggest that the proto-vertebrate SoxE gene possessed both melanogenic and neurogenic capabilities prior to SoxE gene duplication. Following the agnathan-gnathostome split, lamprey SoxE1 and SoxE3 largely lost their melanogenic and/or enteric neurogenic properties, while gnathostome SoxE paralogs have retained functional conservation. We posit that this difference in protein subfunctionalization is a direct consequence of the independent regulation of SoxE paralog expression between the two lineages. Specifically, we propose that the overlapping expression of gnathostome SoxE paralogs in early neural crest largely constrained the function of gnathostome SoxE proteins. In contrast, the largely non-overlapping expression of lamprey SoxE paralogs allowed them to specialize with regard to their DNA-binding and/or protein interaction properties. Restriction of developmental potential among cranial and trunk neural crest in lampreys may be related to constraints on SoxE activity among duplicates, but such specialization does not appear to have occurred in gnathostomes. This highlights an important difference in the evolution of SoxE activity between these two divergent vertebrate lineages and provides insights for understanding how cell fate restriction in different NCC populations may be dependent on subfunctionalization among SoxE duplicates.

Roles for FGF in lamprey pharyngeal pouch formation and skeletogenesis highlight ancestral functions in the vertebrate head.

A defining feature of vertebrates (craniates) is a pronounced head supported and protected by a cellularized endoskeleton. In jawed vertebrates (gnathostomes), the head skeleton is made of rigid three-dimensional elements connected by joints. By contrast, the head skeleton of modern jawless vertebrates (agnathans) consists of thin rods of flexible cellular cartilage, a condition thought to reflect the ancestral vertebrate state. To better understand the origin and evolution of the gnathostome head skeleton, we have been analyzing head skeleton development in the agnathan, lamprey. The fibroblast growth factors FGF3 and FGF8 have various roles during head development in jawed vertebrates, including pharyngeal pouch morphogenesis, patterning of the oral skeleton and chondrogenesis. We isolated lamprey homologs of FGF3, FGF8 and FGF receptors and asked whether these functions are ancestral features of vertebrate development or gnathostome novelties. Using gene expression and pharmacological agents, we found that proper formation of the lamprey head skeleton requires two phases of FGF signaling: an early phase during which FGFs drive pharyngeal pouch formation, and a later phase when they directly regulate skeletal differentiation and patterning. In the context of gene expression and functional studies in gnathostomes, our results suggest that these roles for FGFs arose in the first vertebrates and that the evolution of the jaw and gnathostome cellular cartilage was driven by changes developmentally downstream from pharyngeal FGF signaling.

Reconstructing the ancestral vertebrate brain.

Highly complicated morphologies and sophisticated functions of vertebrate brains have been established through evolution. However, the origin and early evolutionary history of the brain remain elusive, owing to lack of information regarding the brain architecture of extant and fossil species of jawless vertebrates (agnathans). Comparative analyses of the brain of less studied cyclostomes (only extant agnathan group, consisting of lampreys and hagfish) with the well-known sister group of jawed vertebrates (gnathostomes) are the only tools we have available to illustrate the ancestral architecture of the vertebrate brain. Previous developmental studies had shown that the lamprey lacked well-established brain compartments that are present in gnathostomes, such as the medial ganglionic eminence and the rhombic lip. The most accepted scenario suggested that cyclostomes had fewer compartments than that of the gnathostome brain and that gnathostomes thus evolved by a stepwise addition of innovations on its developmental sequence. However, recent studies have revealed that these compartments are present in hagfish embryos, indicating that these brain regions have been acquired before the split of cyclostomes and gnathostomes. By comparing two cyclostome lineages and gnathostomes, it has become possible to speculate about a more complex ancestral state of the brain, excluding derived traits in either of the lineages. In this review, we summarize recent studies on the brain development of the lamprey and hagfish. Then, we attempt to reconstruct the possible brain architecture of the last common ancestor of vertebrates. Finally, we discuss how the developmental plan of the vertebrate brain has been modified independently in different vertebrate lineages.

Evolution of the Vertebrate Cranium: Viewed from Hagfish Developmental Studies.

Our knowledge of vertebrate cranium evolution has relied largely on the study of gnathostomes. Recent evolutionary and developmental studies of cyclostomes have shed new light on the history of the vertebrate skull. The recent ability to obtain embryos of the hagfish, Eptatretus burgeri, has enabled new studies which have suggested an embryonic morphological pattern (the "cyclostome pattern") of craniofacial development. This pattern is shared by cyclostomes, but not by modern jawed vertebrates. Because this pattern of embryonic head development is thought to be present in some stem gnathostomes (ostracoderms), it is possible that the cyclostome pattern represents the vertebrate ancestral pattern. The study of cyclostomes may thus lead to an understanding of the most ancestral basis of craniofacial development. In this review, we summarize the development of the hagfish chondrocranium in light of the cyclostome pattern, present an updated comparison of the cyclostome chondrocranium, and discuss several aspects of the evolution and development of the vertebrate skull.

Body wall development in lamprey and a new perspective on the origin of vertebrate paired fins.

Classical hypotheses regarding the evolutionary origin of paired appendages propose transformation of precursor structures (gill arches and lateral fin folds) into paired fins. During development, gnathostome paired appendages form as outgrowths of body wall somatopleure, a tissue composed of somatic lateral plate mesoderm (LPM) and overlying ectoderm. In amniotes, LPM contributes connective tissue to abaxial musculature and forms ventrolateral dermis of the interlimb body wall. The phylogenetic distribution of this character is uncertain because lineage analyses of LPM have not been generated in anamniotes. We focus on the evolutionary history of the somatopleure to gain insight into the tissue context in which paired fins first appeared. Lampreys diverged from other vertebrates before the acquisition of paired fins and provide a model for investigating the preappendicular condition. We present vital dye fate maps that suggest the somatopleure is eliminated in lamprey as the LPM is separated from the ectoderm and sequestered to the coelomic linings during myotome extension. We also examine the distribution of postcranial mesoderm in catshark and axolotl. In contrast to lamprey, our findings support an LPM contribution to the trunk body wall of these taxa, which is similar to published data for amniotes. Collectively, these data lead us to hypothesize that a persistent somatopleure in the lateral body wall is a gnathostome synapomorphy, and the redistribution of LPM was a key step in generating the novel developmental module that ultimately produced paired fins. These embryological criteria can refocus arguments on paired fin origins and generate hypotheses testable by comparative studies on the source, sequence, and extent of genetic redeployment.

Expression patterns of Eph genes in the "dual visual development" of the lamprey and their significance in the evolution of vision in vertebrates.

Image-forming vision is crucial to animals for recognizing objects in their environment. In vertebrates, this type of vision is achieved with paired camera eyes and topographic projection of the optic nerve. Topographic projection is established by an orthogonal gradient of axon guidance molecules, such as Ephs. To explore the evolution of image-forming vision in vertebrates, lampreys, which belong to the basal lineage of vertebrates, are key animals because they show unique "dual visual development." In the embryonic and pre-ammocoete larval stage (the "primary" phase), photoreceptive "ocellus-like" eyes develop, but there is no retinotectal optic nerve projection. In the late ammocoete larval stage (the "secondary" phase), the eyes grow and form into camera eyes, and retinotectal projection is newly formed. After metamorphosis, this retinotectal projection in adult lampreys is topographic, similar to that of gnathostomes. In this study, we explored the involvement of Ephs in lamprey "dual visual development" and establishment of the image-form vision. We found that gnathostome-like orthogonal gradient expression was present in the retina during the "secondary" phase; i.e., EphB showed a gradient of expression along the dorsoventral axis, while EphC was expressed along the anteroposterior axis. However, no orthogonal gradient expression was observed during the "primary" phase. These observations suggest that Ephs are likely recruited de novo for the guidance of topographical "second" optic nerve projection. Transformations during lamprey "dual visual development" may represent "recapitulation" from a protochordate-like ancestor to a gnathostome-like vertebrate ancestor.

Novel Tfap2-mediated control of soxE expression facilitated the evolutionary emergence of the neural crest.

Gene duplication has been proposed to drive the evolution of novel morphologies. After gene duplication, it is unclear whether changes in the resulting paralogs' coding-regions, or in their cis-regulatory elements, contribute most significantly to the assembly of novel gene regulatory networks. The Transcription Factor Activator Protein 2 (Tfap2) was duplicated in the chordate lineage and is essential for development of the neural crest, a tissue that emerged with vertebrates. Using a tfap2-depleted zebrafish background, we test the ability of available gnathostome, agnathan, cephalochordate and insect tfap2 paralogs to drive neural crest development. With the exception of tfap2d (lamprey and zebrafish), all are able to do so. Together with expression analyses, these results indicate that sub-functionalization has occurred among Tfap2 paralogs, but that neo-functionalization of the Tfap2 protein did not drive the emergence of the neural crest. We investigate whether acquisition of novel target genes for Tfap2 might have done so. We show that in neural crest cells Tfap2 directly activates expression of sox10, which encodes a transcription factor essential for neural crest development. The appearance of this regulatory interaction is likely to have coincided with that of the neural crest, because AP2 and SoxE are not co-expressed in amphioxus, and because neural crest enhancers are not detected proximal to amphioxus soxE. We find that sox10 has limited ability to restore the neural crest in Tfap2-deficient embryos. Together, these results show that mutations resulting in novel Tfap2-mediated regulation of sox10 and other targets contributed to the evolution of the neural crest.

The lamprey: a jawless vertebrate model system for examining origin of the neural crest and other vertebrate traits.

Lampreys are a group of jawless fishes that serve as an important point of comparison for studies of vertebrate evolution. Lampreys and hagfishes are agnathan fishes, the cyclostomes, which sit at a crucial phylogenetic position as the only living sister group of the jawed vertebrates. Comparisons between cyclostomes and jawed vertebrates can help identify shared derived (i.e. synapomorphic) traits that might have been inherited from ancestral early vertebrates, if unlikely to have arisen convergently by chance. One example of a uniquely vertebrate trait is the neural crest, an embryonic tissue that produces many cell types crucial to vertebrate features, such as the craniofacial skeleton, pigmentation of the skin, and much of the peripheral nervous system (Gans and Northcutt, 1983). Invertebrate chordates arguably lack unambiguous neural crest homologs, yet have cells with some similarities, making comparisons with lampreys and jawed vertebrates essential for inferring characteristics of development in early vertebrates, and how they may have evolved from nonvertebrate chordates. Here we review recent research on cyclostome neural crest development, including research on lamprey gene regulatory networks and differentiated neural crest fates.