The TRP channel PKD2 is involved in sensing the mechanical stimulus of adhesion for initiating metamorphosis in the chordate Ciona

Abstract

Metamorphosis is the dramatic and irreversible reconstruction of animal bodies transitioning from the larval stage. Because of the significant impact of metamorphosis on animal life, its timing is strictly regulated. Invertebrate chordate ascidians are the closest living relatives of vertebrates. Ascidians exhibit metamorphosis that converts their swimming larvae into sessile adults. Ascidian metamorphosis is triggered by a mechanical stimulus generated when adhesive papillae adhere to a substrate. However, it is not well understood how the mechanical stimulus is generated and how ascidian larvae sense the stimulus. In this study, we addressed these issues by a combination of embryological, molecular, and genetic experiments in the model ascidian Ciona intestinalis Type A, also called Ciona robusta. We here showed that the epidermal neuronal network starting from the sensory neurons at the adhesive papillae is responsible for the sensing of adhesion. We also found that the transient receptor potential (TRP) channel PKD2 is involved in sensing the stimulus of adhesion. Our results provide a better understanding of the mechanisms underlying the regulation of the timing of ascidian metamorphosis.

Specification of distinct cell types in a sensory-adhesive organ for metamorphosis in theCionalarva

“…This would mirror the development of the epidermal neurons of the tail, in which neurons are born side-by-side as pairs after a final cell division by a committed mother cell (Pasini et al, 2006). Papilla-specific CRISPR knockout of Pou4 with new sgRNAs (Supplemental Figure 3) recapitulated the loss of PN differentiation by the previously published TALEN knockout (Sakamoto et al, 2022), as assayed by C4.78 reporter expression (Figure 6H,I). In contrast, Pou4 knockout had no effect on the specification of ACCs or OCs, suggesting Pou4 function is specific for PN fate in the papillae (Supplemental Figure 4).…”

“…intestinalis Type A) whole-larva single-cell RNA sequencing (scRNAseq) data (Cao et al, 2019) for evidence of the cell types described by transmission electron microscopy (TEM) of the papillae (Zeng et al, 2019b). While a cell cluster annotated as "Palp Sensory Cells" (PSCs) appeared enriched for known markers of ACCs like CryBG (KH.S605.3) and KH.C3.516 (Sharma et al, 2019;Shimeld et al, 2005), genes expressed in other papilla cell types were also enriched in this cluster as well, including Sp6/7/8 (KH.C13.22) (Liu and Satou, 2019;Wagner et al, 2014) and Pou4 (KH.C2.42) (Roure et al, 2022;Sakamoto et al, 2022). Re-analysis and reclustering of these data revealed novel potential markers for these different cell types within the cluster (Supplemental Figure 1A-C, Supplemental Table 1).…”

“…Previous papilla-specific TALEN knockout of the neuronal transcription factor-encoding gene Pou4 successfully eliminated PNs and the larva's tail resorption response to mechanical stimuli (Sakamoto et al, 2022). Pou4 has been previously implicated in a Myt1-dependent regulatory cascade that specifies the caudal epidermal neurons (CENs) of the tail, from neurogenic midline cells expressing the proneural bHLH transcription factor Ascl.a (KH.L9.13, sometimes called Ascl2 or Ascl.b previously) (Pasini et al, 2006;Roure et al, 2022;Roure and Darras, 2016;Tang et al, 2013;Waki et al, 2015).…”

“…With our different CRISPR knockouts affecting different cell types of the papillae, we asked how these different perturbations might affect larval metamorphosis. Only the involvement of the PNs in triggering metamorphosis has been demonstrated (Sakamoto et al, 2022;Wakai et al, 2021), but it is not yet known how the regulatory networks and cell types of the papillae affect different processes during metamorphosis. We performed papilla-specific CRISPR as above using the Foxc>Cas9 vector, targeting the four different transcription factors we have shown to be involved in patterning the cell types of the papillae: Pou4, Islet, Foxg, and Sp6/7/8.…”

“…Several cells appear to secrete the "glue" or bioadhesive material required for the attachment of the larva to the substrate, termed "collocytes" (Zeng et al, 2019a;Zeng et al, 2019b). Other cells are clearly neuronal (four ciliated neurons per papilla) (Zeng et al, 2019b) and are required to trigger the onset of metamorphosis (Sakamoto et al, 2022), which was also recently shown to depend on mechanical stimulation of the papillae (Wakai et al, 2021). Finally, at the very center of each papilla are four "Axial Columnar Cells" (ACCs), which have been suggested to possess chemosensory and contractile properties (Poncelet and Shimeld, 2020;Poncelet et al, 2022;Turon, 1991).…”