Tail regression induced by elevated retinoic acid signaling in amphioxus larvae occurs by tissue remodeling, not cell death

Abstract

The vitamin A derived morphogen retinoic acid (RA) is known to function in the regulation of tissue proliferation and differentiation. Here, we show that exogenous RA applied to late larvae of the invertebrate chordate amphioxus can reverse some differentiated states. Although treatment with the RA antagonist BMS009 has no obvious effect on late larvae of amphioxus, administration of excess RA alters the morphology of the posterior end of the body. The anus closes over, and gut contents accumulate in the hindgut. In addition, the larval tail fin regresses, although little apoptosis takes place. This fin normally consists of columnar epidermal cells, each characterized by a ciliary rootlet running all the way from an apical centriole to the base of the cell and likely contributing substantial cytoskeletal support. After a few days of RA treatment, the rootlet becomes disrupted, and the cell shape changes from columnar to cuboidal. Transmission electron microscopy (TEM) shows fragments of the rootlet in the basal cytoplasm of the cuboidal cell. A major component of the ciliary rootlet in amphioxus is the protein Rootletin, which is encoded by a single AmphiRootletin gene. This gene is highly expressed in the tail epithelial cells of control larvae, but becomes downregulated after about a day of RA treatment, and the breakup of the ciliary rootlet soon follows. The effect of excess RA on these epidermal cells of the larval tail in amphioxus is unlike posterior regression in developing zebrafish, where elevated RA signaling alters connective tissues of mesodermal origin. In contrast, however, the RA-induced closure of the amphioxus anus has parallels in the RA-induced caudal regression syndrome of mammals.

Lineage-specific duplication of amphioxus retinoic acid degrading enzymes (CYP26) resulted in sub-functionalization of patterning and homeostatic roles

“…Previous studies in the Florida amphioxus, B. floridae , have shown that the administration of excess RA during gastrulation results in a small anus [39] and that the continuous administration of RA to B. floridae larvae leads to closure of the anus and regression of the tail fin [51]. Our data on the European amphioxus, B. lanceolatum , are consistent with these previous findings and further suggest that RA signaling is required for proper tail fin outgrowth.…”

“…This downregulation in turn is likely responsible for the induction of tail fin regression upon RA treatments in B. floridae larvae [51]. Our results of RA and CYP26 inhibitor treatments in B. lanceolatum are generally consistent with these previous observations, although we identified a major difference in the developmental timing of the involvement of RA signaling in tail fin outgrowth between the two amphioxus species.…”

“…Given the effects on early tail fin formation we observed in B. lanceolatum in response to the alteration of endogenous RA signaling levels, we decided to investigate the patterns of Rootletin expression in 60-h B. lanceolatum larvae following the pharmacological treatment regimes detailed above. As previously described for late B. floridae larvae [51], Rootletin is expressed in the basal compartment of the columnar tail fin cells in 60-h B. lanceolatum larvae (Fig. 3h).…”

“…It has previously been shown that the amphioxus larval tail fin is composed of columnar epidermal cells that contain a large ciliary rootlet [49, 50] and that RA signaling promotes tail regression in late, pre-metamorphic B. floridae larvae by downregulating the gene encoding the main component of the ciliary rootlet: the protein Rootletin [51]. However, the regulation of Rootletin expression by RA signaling in the tail fin of early B. floridae larvae has not yet been reported.…”

“…The amphioxus tail fin is established by columnar epidermal cells that contain a large ciliary rootlet [49, 50] and it has previously been shown that RA promotes the downregulation of a major component of this structure, the protein Rootletin [51]. This downregulation in turn is likely responsible for the induction of tail fin regression upon RA treatments in B. floridae larvae [51].…”