Abstract
The tunicate Ciona robusta is an emerging model system to study the evolution of the nervous system. Due to their small embryos and compact genomes, tunicates, like Ciona robusta, have great potential to comprehend genetic circuitry underlying cell specific gene repertoire, among different neuronal cells. Their simple larvae possess a sensory vesicle comprising two pigmented sensory organs, the ocellus and the otolith. We focused here on Klhl21/30, a gene belonging to Kelch family, that, in Ciona robusta, starts to be expressed in pigmented cell precursors, becoming specifically maintained in the otolith precursor during embryogenesis. Evolutionary analyses demonstrated the conservation of Klhl21/30 in all the chordates. Cis-regulatory analyses and CRISPR/Cas9 mutagenesis of potential upstream factors, revealed that Klhl21/30 expression is controlled by the combined action of three transcription factors, Mitf, Dmrt, and Msx, which are downstream of FGF signaling. The central role of Mitf is consistent with its function as a fundamental regulator of vertebrate pigment cell development. Moreover, our results unraveled a new function for Dmrt and Msx as transcriptional co-activators in the context of the Ciona otolith.
Highlights
Understanding the developmental logics that orchestrate specific gene expression inside the nervous system represents a fascinating challenge in cell and developmental biology
Using tissuespecific CRISPR/Cas9-mediated gene knockouts, we found that Microphthalmia-associated transcription factor gene (Mitf) is central to Kelch-like 21 (Klhl21)/30 expression in the otolith, as it is for many pigmentation markers in the pigment cells of vertebrates (Levy et al, 2006), with Msx and Dmrt acting as coactivators
We found a Kelch-like (Klhl) gene family member (Kyoto Hoya gene model KH.L84.23), which exhibited similar expression values of known pigment cell precursors (PCP) markers, including Tyr, Tyrp.a and Rab32/38 and that, by reciprocal BLASTs, is similar to vertebrate Kelch-like 21 (Klhl21) and Kelch-like 30 (Klhl30)
Summary
Understanding the developmental logics that orchestrate specific gene expression inside the nervous system represents a fascinating challenge in cell and developmental biology. The identification of the molecular processes underlying cell specific expression among different neuronal cells is very difficult using vertebrate models, due to the relative complexity of vertebrate embryos and genomes, as well as the numbers of genes involved. The tunicate subphylum is the sister group of vertebrates, forming with them the clade Olfactores (Delsuc et al, 2006). Due to their small, invariant embryos and compact genomes (Berná and Alvarez-Valin, 2014), tunicates have great potential to help to uncover the genetic circuitry regulating chordate-specific mechanisms of neural development.
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