A C-terminally truncated form of ß-catenin acts as a novel regulator of Wnt/ß-catenin signaling in planarians.

ß-Catenin, the core element of the Wnt/ß-catenin pathway, is a multifunctional and evolutionarily conserved protein which performs essential roles in a variety of developmental and homeostatic processes. Despite its crucial roles, the mechanisms that control its context-specific functions in time and space remain largely unknown. The Wnt/ß-catenin pathway has been extensively studied in planarians, flatworms with the ability to regenerate and remodel the whole body, providing a 'whole animal' developmental framework to approach this question. Here we identify a C-terminally truncated ß-catenin (ß-catenin4), generated by gene duplication, that is required for planarian photoreceptor cell specification. Our results indicate that the role of ß-catenin4 is to modulate the activity of ß-catenin1, the planarian ß-catenin involved in Wnt signal transduction in the nucleus, mediated by the transcription factor TCF-2. This inhibitory form of ß-catenin, expressed in specific cell types, would provide a novel mechanism to modulate nuclear ß-catenin signaling levels. Genomic searches and in vitro analysis suggest that the existence of a C-terminally truncated form of ß-catenin could be an evolutionarily conserved mechanism to achieve a fine-tuned regulation of Wnt/ß-catenin signaling in specific cellular contexts.

Small C-terminal Domain Phosphatase 3 Dephosphorylates the Linker Sites of Receptor-regulated Smads (R-Smads) to Ensure Transforming Growth Factor ß (TGFß)-mediated Germ Layer Induction in Xenopus Embryos.

Germ layer induction is one of the earliest events shortly after fertilization that initiates body formation of vertebrate embryos. In Xenopus, the maternally deposited transcriptional factor VegT promotes the expression of zygotic Nodal/Activin ligands that further form a morphogen gradient along the vegetal-animal axis and trigger the induction of the three germ layers. Here we found that SCP3 (small C-terminal domain phosphatase 3) is maternally expressed and vegetally enriched in Xenopus embryos and is essential for the timely induction of germ layers. SCP3 is required for the full activation of Nodal/Activin and bone morphogenetic protein signals and functions via dephosphorylation in the linker regions of receptor-regulated Smads. Consistently, the linker regions of receptor-regulated Smads are heavily phosphorylated in fertilized eggs, and this phosphorylation is gradually removed when embryos approach the midblastula transition. Knockdown of maternal SCP3 attenuates these dephosphorylation events and the activation of Nodal/Activin and bone morphogenetic protein signals after midblastula transition. This study thus suggested that the maternal SCP3 serves as a vegetally enriched, intrinsic factor to ensure a prepared status of Smads for their activation by the upcoming ligands during germ layer induction of Xenopus embryos.

A small scale expression screen identifies tissue specific markers in the Dugesia japonica strain Pek-1.

Freshwater planaria has tremendous capacity to reform the missing part of the body and therefore is considered as one of the most important model organism for regeneration study. At present, Schmidtea mediterranea and Dugesia japonica are the two major species utilized for laboratory manipulations. Dugesia japonica flatworms are widely distributed in the Far East including Cherry Valley region in the north-west area of Beijing, China. We reported here the establishment of an asexual Dugesia japonica strain Pek-1, as a suitable system for regeneration study. Using morphological, karyotypical as well as phylogenetic analyses, we confirmed that these flatworms indeed belonged to Dugesia japonica. We went on to show that the commonly used in situ probes and immunohistochemistry reagents and protocols were applicable to the Pek-1 strain. Using this strain, we carried out small scale analysis on EST, RNAi and gene expression. We identified 193 unique EST sequences and 65 of them had not been reported in planarian. By RNAi analysis, we showed that 48 genes, when down-regulated individually, had no effect on regeneration. Furthermore, we identified 3 groups of tissue specific expressing genes that were useful for cell lineage analysis. We concluded that the Dugesia japonica Pek-1 strain could be another suitable animal model to regeneration research.

Djrho2 is involved in regeneration of visual nerves in Dugesia japonica.

The freshwater planarian is a powerful animal model for studying regeneration and stem cell activity in vivo. During regeneration, stem cells (neoblasts in planarian) migrated to the wounding edge to re-build missing parts of the body. However, proteins involved in regulating cell migration during planarian regeneration have not been studied extensively. Here we report two small GTPase genes (Djrho2 and Djrho3) of Dugesia japonica (strain Pek-1). In situ hybridization results indicated that Djrho2 was expressed throughout the body with the exception of the pharynx region while Djrho3 was specifically expressed along the gastro-vascular system. Djrho2 was largely expressed in neoblasts since its expression was sensitive to X-ray irradiation. In Djrho2-RNAi planarians, smaller anterior blastemas were observed in tail fragments during regeneration. Consistently, defective regeneration of visual nerve was detected by immunostainning with VC-1 antibody. These results suggested that Djrho2 is required for proper anterior regeneration in planairan. In contrast, no abnormality was observed after RNAi of Djrho3. We compared protein compositions of control and Djrho2-RNAi planarians using an optimized proteomic approach. Twenty-two up-regulated and 26 de-regulated protein spots were observed in the two-dimensional electrophoresis gels, and 17 proteins were successfully identified by Mass Spectrometry (MS) analysis. Among them, 6 actin-binding or cytoskeleton-related proteins were found de-expressed in Djrho2-RNAi animals, suggesting that abnormal cytoskeleton assembling and cell migration were likely reasons of defected regeneration.