Tissue Regeneration and Biomineralization in Sea Urchins: Role of Notch Signaling and Presence of Stem Cell Markers

Abstract

Echinoderms represent a phylum with exceptional regenerative capabilities that can reconstruct both external appendages and internal organs. Mechanistic understanding of the cellular pathways involved in regeneration in these animals has been hampered by the limited genomic tools and limited ability to manipulate regenerative processes. We present a functional assay to investigate mechanisms of tissue regeneration and biomineralization by measuring the regrowth of amputated tube feet (sensory and motor appendages) and spines in the sea urchin, Lytechinus variegatus. The ability to manipulate regeneration was demonstrated by concentration-dependent inhibition of regrowth of spines and tube feet by treatment with the mitotic inhibitor, vincristine. Treatment with the gamma-secretase inhibitor DAPT resulted in a concentration-dependent inhibition of regrowth, indicating that both tube feet and spine regeneration require functional Notch signaling. Stem cell markers (Piwi and Vasa) were expressed in tube feet and spine tissue, and Vasa-positive cells were localized throughout the epidermis of tube feet by immunohistochemistry, suggesting the existence of multipotent progenitor cells in these highly regenerative appendages. The presence of Vasa protein in other somatic tissues (e.g. esophagus, radial nerve, and a sub-population of coelomocytes) suggests that multipotent cells are present throughout adult sea urchins and may contribute to normal homeostasis in addition to regeneration. Mechanistic insight into the cellular pathways governing the tremendous regenerative capacity of echinoderms may reveal processes that can be modulated for regenerative therapies, shed light on the evolution of regeneration, and enable the ability to predict how these processes will respond to changing environmental conditions.

Active Notch Signaling is Required for Arm Regeneration in a Brittle Star

“…A similar effect of Notch pathway inhibition on regeneration was previously shown for sea urchin spines and podia. Treatment of DAPT resulted in a dosage-dependent reduction of regrowth of those structures post-amputation (20). This suggests that the involvement of Notch signaling in the regeneration of body appendages is conserved among echinoderm classes.…”

“…The only context, in which expression of those genes was studied at the cell and tissue levels in echinoderms, was sea urchin embryogenesis (18,19). The only functional study of the Notch signaling pathway in the context of adult echinoderm regeneration was performed in the sea urchin Lytechinus variegatus (20). This work demonstrated the requirement of the functional Notch signaling for the proper outgrowth of amputated external appendages, such as spines and podia of Lytechinus variegatus (Lamarck, 1816).…”

Germ Line Mechanics—And Unfinished Business

“…The sea urchin embryo was of particular interest. Vasa expression was present throughout the early embryo, in cells for example, that gave rise only to somatic ectoderm, or in larval tissues of the ectoderm (Yajima and Wessel, 2015), and recently reported even in tissues of the adult tube feet (Reinardy et al, 2015). Moreover, when Vasa expression was knocked-down in the embryos, the cell cycle of those embryos was highly disorganized (Yajima and Wessel, 2011).…”

Anterior regeneration after fission in the holothurianCladolabes schmeltzii(Dendrochirotida: Holothuroidea)

“…It is unclear what cells give rise to primordial germ cells in adult animals. Theoretically, three sources can be assumed: stem cells that are probably still present in echinoderms (Candia Carnevali et al, 2009;Reinardy et al, 2015); transdifferentiation of specialized cells of tissue remnants; migration of primordial germ cells into the posterior fragment prior to fission. Kille (1942) did not find migrating germ cells, but, according to his supposition, primordial germ Stored nutrients play a major role in regeneration, because animals cannot feed when damaged.…”