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

University of Washington Seattle, Seattle, Washington, United States
Developmental Biology (Impact Factor: 3.55). 07/2007; 306(1):170-8. DOI: 10.1016/j.ydbio.2007.03.014
Source: PubMed


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.

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    • "To detect transcripts on sectioned samples, in situ hybridization on frozen sections was carried out as previously described (Ohgo et al., 2010) with slight modification. To synthesize an antisense RNA probe for gfp, CMV-GFP5 plasmid (Yokoyama et al., 2007) was linearized with BamHI and transcribed with T7 RNA polymerase (Roche). Probes for prrx1 (Suzuki et al., 2005) and dusp6 (Gomez et al., 2005) were synthesized as previously described. "
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    ABSTRACT: Many amphibians can regenerate limbs, even in adulthood. If a limb is amputated, the stump generates a blastema that makes a complete, new limb in a process similar to developmental morphogenesis. The blastema is thought to inherit its limb-patterning properties from cells in the stump, and it retains the information despite changes in morphology, gene expression, and differentiation states required by limb regeneration. We hypothesized that these cellular properties are maintained as epigenetic memory through histone modifications. To test this hypothesis, we analyzed genome-wide histone modifications in Xenopus limb bud regeneration. The trimethylation of histone H3 at lysine 4 (H3K4me3) is closely related to an open chromatin structure that allows transcription factors access to genes, whereas the trimethylation of histone H3 at lysine 27 (H3K27me3) is related to a closed chromatin state that blocks the access of transcription factors. We compared these two modification profiles by high-throughput sequencing of samples prepared from the intact limb bud and the regenerative blastema by chromatin immunoprecipitation. For many developmental genes, histone modifications at the transcription start site were the same in the limb bud and the blastema, were stable during regeneration, and corresponded well to limb properties. These results support our hypothesis that histone modifications function as a heritable cellular memory to maintain limb cell properties, despite dynamic changes in gene expression during limb bud regeneration in Xenopus. Copyright © 2015. Published by Elsevier Inc.
    Developmental Biology 08/2015; DOI:10.1016/j.ydbio.2015.08.013 · 3.55 Impact Factor
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    • "Inhibition of Wnt/ß-catenin signaling in the regenerating tail fin or limb bud blocks regeneration (Wehner et al., 2014; Yokoyama et al., 2007), suggesting that the pathway may be Fig. 3. Representative images of gfap:Cre ERT2 -labeled progeny in the regenerating blastema. (A–C) At 5 dpi one mCherry þ cell is GFAP þ (arrow), while a neighboring cell is GFAP À (arrowhead). "
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    Developmental Biology 04/2015; 403(1). DOI:10.1016/j.ydbio.2015.03.025 · 3.55 Impact Factor
    • "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. "
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