An Amphioxus Gli Gene Reveals Conservation of Midline Patterning and the Evolution of Hedgehog Signalling Diversity in Chordates

Background. Hedgehog signalling, interpreted in receiving cells by Gli transcription factors, plays a central role in the development of vertebrate and Drosphila embryos. Many aspects of the signalling pathway are conserved between these lineages, however vertebrates have diverged in at least one key aspect: they have evolved multiple Gli genes encoding functionally-distinct proteins, increasing the complexity of the hedgehog-dependent transcriptional response. Amphioxus is one of the closest living relatives of the vertebrates, having split from the vertebrate lineage prior to the widespread gene duplication prominent in early vertebrate evolution. Principal findings. We show that amphioxus has a single Gli gene, which is deployed in tissues adjacent to sources of hedgehog signalling derived from the midline and anterior endoderm. This shows the duplication and divergence of the Gli family, and hence the origin of vertebrate Gli functional diversity, was specific to the vertebrate lineage. However we also show that the single amphioxus Gli gene produces two distinct transcripts encoding different proteins. We utilise three tests of Gli function to examine the transcription regulatory capacities of these different proteins, demonstrating one has activating activity similar to Gli2, while the other acts as a weak repressor, similar to Gli3. Conclusions. These data show that the vertebrates and amphioxus have evolved functionally-similar repertoires of Gli proteins using parallel molecular routes; vertebrates via gene duplication and divergence, and amphioxus via alternate splicing of a single gene. Our results demonstrate that similar functional complexity of intercellular signalling can be achieved via different evolutionary pathways

Molecular regionalization of the developing amphioxus neural tube challenges major partitions of the vertebrate brain

All vertebrate brains develop following a common Bauplan defined by anteroposterior (AP) and dorsoventral (DV) subdivisions, characterized by largely conserved differential expression of gene markers. However, it is still unclear how this Bauplan originated during evolution. We studied the relative expression of 48 genes with key roles in vertebrate neural patterning in a representative amphioxus embryonic stage. Unlike nonchordates, amphioxus develops its central nervous system (CNS) from a neural plate that is homologous to that of vertebrates, allowing direct topological comparisons. The resulting genoarchitectonic model revealed that the amphioxus incipient neural tube is unexpectedly complex, consisting of several AP and DV molecular partitions. Strikingly, comparison with vertebrates indicates that the vertebrate thalamus, pretectum, and midbrain domains jointly correspond to a single amphioxus region, which we termed Di-Mesencephalic primordium (DiMes). This suggests that these domains have a common developmental and evolutionary origin, as supported by functional experiments manipulating secondary organizers in zebrafish and mice.Spanish Ministry of Economy and Competitiveness and European FEDER funds (grant number BFU2014-57516-P). To Luis Puelles and Jose Luis Ferran. European Research Council (grant number ERC-StG-LS2-637591). To Manuel Irimia. Spanish Ministry of Economy and Competitiveness (grant number SEV-2012-0208).Centro de Excelencia Severo Ochoa (to CRG, Manuel Irimia). Spanish Ministry of Economy and Competitiveness (grant number BFU2014-58908-P). To Jordi Garcia-Fernadez. Seneca Foundation, Comunidad de Murcia (grant number 19904/GERM/15). To Luis Puelles. Generalitat de Catalunya (grant number). ICREA Academia Prize to Jordi Garcia-Fernandez. Spanish Ministry of Economy and Competitiveness (grant number BFU2013-43213-P). To Paola Bovolenta. Spanish Ministry of Economy and Competitiveness (grant number BFU2014-55076-P). To Manuel Irimia. Including an FPI PhD fellowship to Laura Lopez-Blanch. Marine Alliance for Science and Technology Scotland (MASTS) (grant number). To Ildiko Somorjai