Wnt/β-catenin signaling has an essential role in the initiation of limb regeneration

Abstract

Anuran (frog) tadpoles and urodeles (newts and salamanders) are the only vertebrates capable of fully regenerating amputated limbs. During the early stages of regeneration these amphibians form a "blastema", a group of mesenchymal progenitor cells that specifically directs the regrowth of the limb. We report that wnt-3a is expressed in the apical epithelium of regenerating Xenopus laevis limb buds, at the appropriate time and place to play a role during blastema formation. To test whether Wnt/beta-catenin signaling is required for limb regeneration, we created transgenic X. laevis tadpoles that express Dickkopf-1 (Dkk1), a specific inhibitor of Wnt/beta-catenin signaling, under the control of a heat-shock promoter. Heat-shock immediately before limb amputation or during early blastema formation blocked limb regeneration but did not affect the development of contralateral, un-amputated limb buds. When the transgenic tadpoles were heat-shocked following the formation of a blastema, however, they retained the ability to regenerate partial hindlimb structures. Furthermore, heat-shock induced Dkk1 blocked fgf-8 but not fgf-10 expression in the blastema. We conclude that Wnt/beta-catenin signaling has an essential role during the early stages of limb regeneration, but is not absolutely required after blastema formation.

Beyond early development: Xenopus as an emerging model for the study of regenerative mechanisms

“…However, in the limb, similar circumstantial expression data also places Wnt signaling upstream of Fgf (Yokoyama et al, 2007). Brief induction of an HS-Dkk1 transgene on day 5 of limb regeneration abolished fgf8 expression 8 hr later, while fgf10, lmx1, msx2, and hoxa13 expression are unaltered.…”

“…The transgenic method combined with the heat inducible promoter hsp70 (HS) is a powerful tool to manipulate specific pathways at a specific time during the regeneration process. It has been used many times in Xenopus and zebrafish to dissect a specific molecular pathway used during regeneration (see Section 2.2; Beck et al, 2003Beck et al, , 2006Lee et al, 2005;Stoick-Cooper et al, 2007b;Yokoyama et al, 2007;Lin and Slack, 2008). To direct the spatial action of a protein more precisely, beads soaked in recombinant protein solution or cell aggregates expressing a specific protein have been grafted into the amputation stump Satoh et al, 2005a;Sugiura et al, 2008) and virus infection has also been used (Kawakami et al, 2006).…”

“…In the past few years, several studies demonstrated a role for the Wnt/␤-catenin signaling pathway during regeneration (Kawakami et al, 2006;Yokoyama et al, 2007;Lin and Slack, 2008;Sugiura et al, 2008). Wnt signaling can be blocked by over-expressing the secreted antagonist dickkopf-1 (Dkk-1) using a heat inducible transgene (HS-dkk-1).…”

Fine-mapping quantitative trait loci affecting murine external ear tissue regeneration in the LG/J by SM/J advanced intercross line

“…Also, WNT proteins regulate the rate of satellite cell proliferation during adult skeletal muscle cell regeneration (Otto et al, 2008). WNT has been shown to have a key role in the initiation of amphibian limb (Kawakami et al, 2006;Yokoyama et al, 2007) and zebrafish fin regeneration (Goessling et al, 2009). Wnt3a has only one highly conserved intronic noncoding SNP in its sequence.…”

Localization of planarian β-CATENIN-1 reveals multiple roles during anterior-posterior regeneration and organogenesis

“…The canonical (β-catenin-dependent) Wnt pathway is an evolutionarily conserved signaling pathway in animals which controls multiple crucial events during development, adult homeostasis and body regeneration (Clevers et al, 2014;Huelsken and Birchmeier, 2001;Martin and Kimelman, 2009;van Amerongen and Nusse, 2009;Yokoyama et al, 2007). During canonical signaling, secreted Wnt ligands bind to their Frizzled receptors to induce cytoplasmic accumulation and nuclear translocation of β-catenin (Clevers and Nusse, 2012;Mikels and Nusse, 2006), which is the key intracellular component of the transduction cascade.…”