Apoptosis is a generator of Wnt-dependent regeneration and homeostatic cell renewal in the ascidian Ciona

Abstract

Body regeneration is unilateral in the ascidian Ciona intestinalis: severed basal body parts can regenerate distal structures, such as the siphons and neural complex, but severed distal body parts do not replace basal structures. Regeneration involves the activity of adult stem cells in vasculature of the branchial sac, which are induced to proliferate and produce migratory progenitor cells for the replacement of missing tissues and organs. Branchial sac-derived stem cells also replenish continuously recycling cells lining the pharyngeal fissures during homeostatic growth. Apoptosis at injury sites is an early and transient event of regeneration and occurs continuously in the pharyngeal fissures during homeostatic growth. Treatment of amputated animals with caspase 1 inhibitor, caspase 3 inhibitor, or the pan-caspase inhibitor Z-VAD-FMK blocked apoptosis, prevented regeneration, and suppressed the growth and function of the branchial sac. A pharmacological screen and inhibitory siRNA treatments indicated that regeneration and homeostatic growth require canonical Wnt signaling. Furthermore, exogenously supplied recombinant Wnt3a protein rescued both caspase-blocked regeneration and normal branchial sac growth. As determined by EdU pulse-chase studies, inhibition of apoptosis did not affect branchial sac stem cell proliferation but instead prevented the survival of progenitor cells. After bisection across the mid-body, apoptosis at the injury site occurred in the regenerating basal fragments, but not in the non-regenerating distal fragments, although both fragments contain a large portion of the branchial sac, suggesting that apoptosis is unilateral at the wound site and the presence of branchial sac stem cells is insufficient for regeneration. The results show that apoptosis-dependent Wnt signaling mediates regeneration and homeostatic growth by promoting progenitor cell survival in Ciona.