Neural crest cells (NCCs) are migratory, multipotent embryonic cells that are unique to vertebrates and form an array of clade-defining adult features. The evolution of NCCs has been linked to ...various genomic events, including the evolution of new gene-regulatory networks
, the de novo evolution of genes
and the proliferation of paralogous genes during genome-wide duplication events
. However, conclusive functional evidence linking new and/or duplicated genes to NCC evolution is lacking. Endothelin ligands (Edns) and endothelin receptors (Ednrs) are unique to vertebrates
, and regulate multiple aspects of NCC development in jawed vertebrates
. Here, to test whether the evolution of Edn signalling was a driver of NCC evolution, we used CRISPR-Cas9 mutagenesis
to disrupt edn, ednr and dlx genes in the sea lamprey, Petromyzon marinus. Lampreys are jawless fishes that last shared a common ancestor with modern jawed vertebrates around 500 million years ago
. Thus, comparisons between lampreys and gnathostomes can identify deeply conserved and evolutionarily flexible features of vertebrate development. Using the frog Xenopus laevis to expand gnathostome phylogenetic representation and facilitate side-by-side analyses, we identify ancient and lineage-specific roles for Edn signalling. These findings suggest that Edn signalling was activated in NCCs before duplication of the vertebrate genome. Then, after one or more genome-wide duplications in the vertebrate stem, paralogous Edn pathways functionally diverged, resulting in NCC subpopulations with different Edn signalling requirements. We posit that this new developmental modularity facilitated the independent evolution of NCC derivatives in stem vertebrates. Consistent with this, differences in Edn pathway targets are associated with differences in the oropharyngeal skeleton and autonomic nervous system of lampreys and modern gnathostomes. In summary, our work provides functional genetic evidence linking the origin and duplication of new vertebrate genes with the stepwise evolution of a defining vertebrate novelty.
Lamprey is one of only two living jawless vertebrates, a group that includes the first vertebrates. Comparisons between lamprey and jawed vertebrates have yielded important insights into the origin ...and evolution of vertebrate physiology, morphology and development. Despite its key phylogenetic position, studies of lamprey have been limited by their complex life history, which makes traditional genetic approaches impossible. The CRISPR/Cas9 system is a bacterial defense mechanism that was recently adapted to achieve high-efficiency targeted mutagenesis in eukaryotes. Here we report CRISPR/Cas9-mediated disruption of the genes Tyrosinase and FGF8/17/18 in the sea lamprey Petromyzon marinus, and detail optimized parameters for producing mutant F0 embryos. Using phenotype and genotype analyses, we show that CRISPR/Cas9 is highly effective in the sea lamprey, with a majority of injected embryos developing into complete or partial mutants. The ability to create large numbers of mutant embryos without inbred lines opens exciting new possibilities for studying development in lamprey and other non-traditional model organisms with life histories that prohibit the generation of mutant lines.
The apparent evolvability of the vertebrate head skeleton has allowed a diverse array of shapes, sizes, and compositions of the head in order to better adapt species to their environments. This ...encompasses feeding, breathing, sensing, and communicating: the head skeleton somehow participated in the evolution of all these critical processes for the last 500 million years. Through evolution, present head diversity was made possible via developmental modifications to the first head skeletal genetic program. Understanding the development of the vertebrate common ancestor's head skeleton is thus an important step in identifying how different lineages have respectively achieved their many innovations in the head. To this end, cyclostomes (jawless vertebrates) are extremely useful, having diverged from jawed vertebrates approximately 400 million years ago, at the deepest node within living vertebrates. From this ancestral vantage point (that is, the node connecting cyclostomes and gnathostomes) we can best identify the earliest major differences in development between vertebrate classes, and start to address how these might translate onto morphology. In this review we survey what is currently known about the cell biology and gene expression during head development in modern vertebrates, allowing us to better characterize the developmental genetics driving head skeleton formation in the most recent common ancestor of all living vertebrates. By pairing this vertebrate composite with information from fossil chordates, we can also deduce how gene regulatory modules might have been arranged in the ancestral vertebrate head. Together, we can immediately begin to understand which aspects of head skeletal development are the most conserved, and which are divergent, informing us as to when the first differences appear during development, and thus which pathways or cell types might be involved in generating lineage specific shape and structure.
•Comparing jawed and jawless vertebrates reveals conserved aspects of head development.•3 molecularly distinct streams of cranial neural crest cells arose in the vertebrate stem.•dlx nesting in the pharynx was likely present in the vertebrate common ancestor.•An ancient head skeletal prepattern utilized hox, alx, hand, msx, and prrx genes.•Most lineage specific morphologies likely arise downstream of this prepattern.
The evolution of vertebrates from an invertebrate chordate ancestor involved the evolution of new organs, tissues, and cell types. It was also marked by the origin and duplication of new gene ...families. If, and how, these morphological and genetic innovations are related is an unresolved question in vertebrate evolution. Hyaluronan is an extracellular matrix (ECM) polysaccharide important for water homeostasis and tissue structure. Vertebrates possess a novel family of hyaluronan binding proteins called Lecticans, and studies in jawed vertebrates (gnathostomes) have shown they function in many of the cells and tissues that are unique to vertebrates. This raises the possibility that the origin and/or expansion of this gene family helped drive the evolution of these vertebrate novelties. In order to better understand the evolution of the lectican gene family, and its role in the evolution of vertebrate morphological novelties, we investigated the phylogeny, genomic arrangement, and expression patterns of all lecticans in the sea lamprey (Petromyzon marinus), a jawless vertebrate. Though both P. marinus and gnathostomes each have four lecticans, our phylogenetic and syntenic analyses are most consistent with the independent duplication of one of more lecticans in the lamprey lineage. Despite the likely independent expansion of the lamprey and gnathostome lectican families, we find highly conserved expression of lecticans in vertebrate-specific and mesenchyme-derived tissues. We also find that, unlike gnathostomes, lamprey expresses its lectican paralogs in distinct subpopulations of head skeleton precursors, potentially reflecting an ancestral diversity of skeletal tissue types. Together, these observations suggest that the ancestral pre-duplication lectican had a complex expression pattern, functioned to support mesenchymal histology, and likely played a role in the evolution of vertebrate-specific cell and tissue types.
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•Both P. marinus and gnathostomes have four lecticans, likely resulting from lineage-specific duplications.•Despite the likely independent expansion in both lineages, lectican expression is highly conserved in lampreys and gnathostomes.•Lamprey expresses lectican genes in distinct cartilage populations, possibly reflecting an ancestral set of skeletal tissues.•The ancestral lectican supported mesenchymal histology, and it likely facilitated vertebrate-specific cell type evolution.
4 Neural crest-derived cellular cartilage is one of the defining characteristics of vertebrates. Elaboration of this tissue and its patterning allowed the evolution of jaws in the gnathostome ...lineage. Together these hallmarks helped jawed vertebrates become one of the dominant taxons in the animal kingdom. Lampreys, as basal jawless vertebrates, lie at a unique phylogenetic position that makes them ideal organisms for the study of evolution of vertebrate/gnathostome novelties. Larval lampreys possess a special oral skeleton composed of a tissue related to cartilage, termed mucocartilage. Despite considerable attention that has been paid to the evolutionary significance of mucocartilage, it is not yet clear, how this unique feature arises in development and to what extent it is homologous to gnathostome jaws. In this study, the development of oro-pharyngeal region was analyzed in the sea lamprey Petromyzon marinus. SEM imaging revealed shaping and topographic relationships of embryonic tissues, detailed plastic histology coupled with expression analyses of several molecular markers were used to describe origin, histogenesis and morphogenesis of mucocartilage. Furthermore, genetic regulation of the tissue was investigated in order to identify its unique or shared features. Mucocartilage is seen to...
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