Highly distinct genetic programs for peripheral nervous system formation in chordates

Abstract

Background Vertebrates develop their peripheral nervous system (PNS) from transient unique embryonic structures, the neural crest, and the ectodermal placodes that are located at the border of the forming central nervous system. By contrast, in the invertebrate chordates, amphioxus and ascidians, a large part of the PNS originates at the opposite of the embryo, in the ventral ectoderm. In both groups, a biphasic mechanism regulates ventral PNS formation: high BMP levels specify a neurogenic territory within which glutamatergic epidermal sensory neuron formation is controlled by the Notch pathway. Given these similarities and the phylogenetic relationships within chordates, it is likely that ventral PNS is an ancestral feature in chordates and that it has been lost in vertebrates. Results In order to get insights into the molecular control of ventral PNS formation and to test the hypothesis of their homology and potential contribution to the emergence of vertebrate PNS, we undertook a close comparison of ventral PNS formation in the ascidian Phallusia mammillata and the amphioxus Branchiostoma lanceolatum. Using timed RNA-seq series, we identified novel markers of the ventral PNS during different phases of its development in both species. By extensively determining the expression of paralogous and orthologous genes, we observed that only a minority of genes have a shared expression in the ventral PNS. However, a large fraction of ventral PNS orthologous genes are expressed in the dorsally forming PNS of vertebrates. Conclusions Our work has significantly increased the molecular characterization of ventral PNS formation in invertebrate chordates. The low observed conservation of gene expression in the ventral PNS suggests that the amphioxus and ascidian ventral PNS are either not homologous, or alternatively extensive drift has occurred in their regulatory mechanisms following a long period (600 My) of separate evolution and accelerated evolution in the ascidian lineage. The homology to genes expressed in the dorsally forming PNS of vertebrates suggests that ancestral sensory neurons gene networks have been redeployed in vertebrates.

Regulation of anterior neurectoderm specification and differentiation by BMP signaling in ascidians

“…First, we determined that BMP signaling was active is the ventral part of the embryo with a similar dynamic to Ciona as revealed by P-Smad1/5/8 immunostaining (Fig S2). Next, we identified single orthologs for Celf3/4/5/6, Pou4 and Isl genes, that were all expressed in the palps (Fig 7) (Chowdhury et al, 2022; Coulcher et al, 2020; Dardaillon et al, 2020).…”

“…Given the overall similar effects on palps formation after alterations of BMP signaling, we sought to identify novel palps molecular markers by using a dataset previously generated in P. mammillata (Chowdhury et al, 2022). We had generated, at several developmental stages, RNA-seq data for whole embryos treated with recombinant BMP4 protein and/or DAPT, a pharmacological Notch inhibitor.…”

“…Using Gene Ontology analysis, we selected a list of 53 genes encoding developmental regulators (transcription factors and signaling molecules) or involved in neural tissue formation, and examined their expression patterns (Table S2). By searching the Aniseed database (Dardaillon et al, 2020) and our previously generated expression data (Chowdhury et al, 2022; Coulcher et al, 2020), we identified 12/26 genes expressed in the palps. By performing in situ hybridization for 27 extra genes, we discovered 7 novel palps markers whose expression is shown in Fig 8.…”

“…Surprisingly, by examining the expression data generated previously, we found that some genes with palps expression were up-regulated by BMP in our dataset, such as Chrdl and Nos (Table S2 and Fig 8). To have a broader view of the potential effect of BMP signaling on gene regulation in the palps, we gathered, from previous publications (Chen et al, 2011; Chowdhury et al, 2022; Coulcher et al, 2020; Joyce Tang et al, 2013; Kusakabe et al, 2012; Liu and Satou, 2019; Pasini et al, 2006; Roure and Darras, 2016; Shimeld et al, 2005; Wagner and Levine, 2012; Wagner et al, 2014), from the Aniseed database (Dardaillon et al, 2020) and from the present study, a list of 68 genes with expression in the palps lineage in Ciona and/or Phallusia (Table S3). We plotted the results of our Phallusia RNA-seq data, and found that 70% of the genes were regulated by BMP signaling.…”

“…Interestingly, we show that modulating BMP pathway in the ascidian Phallusia mammillata (275 My of divergence time) produces the same phenotypes as in C. intestinalis . This allowed us to use previously published RNA-seq data (Chowdhury et al, 2022) to identify a number of novel genes expressed in the ANB and the palps. Altogether, our work points to a role for signaling pathways inhibition in ANB specification, similarly to early anterior neurectoderm formation in vertebrates.…”