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Abstract
Although apoptosis has been widely observed during the regenerative process, the mechanisms by which it is regulated and its roles in regeneration remained unclear. In this study, we introduced Aeolosoma viride, a fresh water annelid with an extraordinary regenerative ability as our model organism to study the functions and regulations of apoptotic caspases. Here we showed that major events of apoptosis were detected near the wounded area and showed spatial correlation with the expression patterns of caspase gene namely Avi-caspase X and two apoptosis regulators namely Avi-Bax and Avi-Bcl-xL. Next, we investigated how Avi-caspase X gene expression and apoptosis influence regeneration following head amputation. RNA interference of Avi-caspase X reduced the amounts of apoptotic cells, as well as the percentage of successful regeneration, suggesting a critical role for apoptosis in anterior regeneration of A. viride. In addition, we also discovered that the expression of apoptotic caspases was regulated by the canonical Wnt signaling pathway. Together, our study showed that caspase dependent apoptosis was critical to the anterior regeneration of A. viride, and could be regulated by the canonical Wnt signaling pathway.
Highlights
Apoptosis has been widely observed during the regenerative process, the mechanisms by which it is regulated and its roles in regeneration remained unclear
In order to determine whether apoptosis takes place during the anterior regeneration of A. viride, Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was performed to label the apoptotic and necrotic cells at different stages of anterior regeneration
Apoptotic cells were recognized in yellow, as confirmed by the positive control, and the nuclei were counterstained in red by propidium iodide (PI), as confirmed by the negative control (Fig. S1)
Summary
Apoptosis has been widely observed during the regenerative process, the mechanisms by which it is regulated and its roles in regeneration remained unclear. Our study showed that caspase dependent apoptosis was critical to the anterior regeneration of A. viride, and could be regulated by the canonical Wnt signaling pathway. The association between the canonical Wnt signaling pathway and apoptosis has become more evident in recent literatures, studies have shown that apoptosis served as an unexpected sources of Wnt[3] positive cell in hydra and Wnt[11] induced cardiomyocyte development through caspase mediate suppression[31,32,33]. To further understand the development and the evolutionary aspect of this relationship, our study aims to investigate the association between apoptosis and canonical Wnt signaling pathway in this regenerating annelid species. Avi-wnt-4 was previously identified as a key regulator of canonical Wnt signaling pathway[38] Together these results suggested that gene expression of Avi-caspase X is critical to anterior regeneration of A. viride, and is regulated by the Wnt/β-catenin signaling pathway
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