The tail of the frog tadpole, comprising spinal cord, muscle, and notochord, regenerates following partial amputation. We show that, in Xenopus, this occurs throughout development, except for a "refractory period" between stages 45 and 47, when tails heal over without regeneration. Regeneration can be enabled during this refractory period by activation of either the BMP or Notch signaling pathways. Conversely, regeneration can be prevented during the later, regenerative, stages by inhibition of either pathway. BMP signaling will cause regeneration of all tissues, whereas Notch signaling activates regeneration of spinal cord and notochord, but not muscle. An activated form of Msx1 can promote regeneration in the same way as BMP signaling. Epistasis experiments suggest that BMP signaling is upstream of Notch signaling but exerts an independent effect on muscle regeneration. The results demonstrate that regenerative capability can be enabled by genetic modifications that reactivate specific components of the developmental program.

Early in Drosophila embryogenesis, gap gene products directly repress transcription of homeotic (HOX) genes and thereby delimit HOX expression domains. Subsequently, Polycomb-group proteins maintain this repression. Currently, there is no known molecular link between gap and Polycomb-group proteins. Here, dMi-2 is identified as a protein that binds to a domain in the gap protein Hunchback that is specifically required for the repression of HOX genes. Genetic analyses show that dMi-2 participates in both Hunchback and Polycomb repression in vivo. Hence, recruitment of dMi-2 may serve as a link between repression of HOX genes by Hunchback and Polycomb proteins.

FGF-8 has attracted attention particularly because of its importance for limb development in the chick and mouse, although it also has a number of earlier expression domains in these species. We have now cloned an FGF-8 homologue from Xenopus in which it is easier to do functional studies on early development. There is no maternal expression, while zygotic expression is highest in the gastrula and neurula stages. XFGF-8 is expressed as a ring around the blastopore and subsequently in the tail bud. There are several domains in the head including the hatching gland, the branchial clefts, and the midbrain-hindbrain border. At later stages there is a prominent band of expression in the limb bud epidermis. Although there is no morphological apical ridge, this band of expression suggests that the Xenopus limb bud contains a cryptic region with a similar ability to stimulate mesenchymal outgrowth. The mesoderm-inducing activity of XFGF-8 is somewhat lower than that of other FGFs, while the posteriorizing activity is similar. These differences are probably due to the different receptor specificity. The posterior expression and high posteriorizing activity suggest that XFGF-8 contributes to the patterning of the anterior-posterior axis by FGF family members during gastrulation. In contrast to the amniotes, Xenopus limb buds can regenerate following damage. We show that regeneration is correlated with the reexpression of XFGF-8 in the distal epidermis, suggesting that this ability is critical for successful limb regeneration.

Ligand-independent activation of the sevenless receptor tyrosine kinase changes the fate of cells in the developing Drosophila eye Basler, K; Christen, B; Hafen, E Basler, K; Christen, B; Hafen, E. Ligand-independent activation of the sevenless receptor tyrosine kinase changes the fate of cells in the developing Drosophila eye. Cell 1991, 64(6) Ligand-independent activation of the sevenless receptor tyrosine kinase changes the fate of cells in the developing Drosophila eye AbstractCell fate in the developing eye is determined by a cascade of inductive interactions. In this process, the sevenless protein--a receptor tyrosine kinase--is required for the specification of the R7 photoreceptor cell fate. We have constructed a gain-of-function sevenless mutation (SevS11) by overexpressing a truncated sevenless protein in the cells where sevenless is normally expressed. In SevS11 mutant flies, all sevenless-expressing cells initiate neural development. This results in the formation of multiple R7-like photoreceptors per ommatidium. Therefore, sevenless activity appears to be necessary and sufficient for the determination of R7 cell fate. These results illustrate the central role receptor tyrosine kinases can play in the specification of cell fate during development. In this process, the sevenless protein-a receptor tyrosine kinase-is required for the specification of the R7 photoreceptor cell fate. We have constructed a gain-of-function sexenless mutation (SetP) by overexpressing a truncated sevenless proteln in the ceils where sevenless is normally expressed. In Se+ mutant flies, all sevenless-expressing cells initiate neural development. This results in the formation of multiple RFltke photorecep tors perommatidium.Therefore, sevenless activity ap pears to be necessary and sufficient for the determination of R7 cell fate. These results illustrate the central role receptortyrosine klnases can play in the specification of cell fate during development.

Bone morphogenetic protein (BMP) signalling is necessary for both the development of the tail bud and for tail regeneration in Xenopus laevis tadpoles. Using a stable transgenic line in which expression of the soluble BMP inhibitor noggin is under the control of the temperature inducible hsp70 promoter, we have investigated the timing of the requirement for BMP signalling during tail regeneration. If noggin expression is induced followed by partial amputation of the tail, then wound closure and the formation of the neural ampulla occur normally but outgrowth of the regeneration bud is inhibited. Furthermore, we show that BMP signalling is also necessary for limb bud regeneration, which occurs in Xenopus tadpoles prior to differentiation. When noggin expression is induced, limb bud regeneration fails at an early stage and a stump is formed. The situation appears similar to the tail, with formation of the limb bud blastema occurring but renewed outgrowth inhibited. The transcriptional repressor Msx1, a direct target of BMP signalling with known roles in vertebrate appendage regeneration, fails to be re-expressed in both tail and limb in the presence of noggin. DNA labelling studies show that proliferation in the notochord and spinal cord of the tail, and of the blastema in the limb bud, is significantly inhibited by noggin induction, suggesting that in the context of these regenerating appendages BMP is mainly required, directly or indirectly, as a mitogenic factor.

While Xenopus is a well-known model system for early vertebrate development, in recent years, it has also emerged as a leading model for regeneration research. As an anuran amphibian, Xenopus laevis can regenerate the larval tail and limb by means of the formation of a proliferating blastema, the lens of the eye by transdifferentiation of nearby tissues, and also exhibits a partial regeneration of the postmetamorphic froglet forelimb. With the availability of inducible transgenic techniques for Xenopus, recent experiments are beginning to address the functional role of genes in the process of regeneration. The use of soluble inhibitors has also been very successful in this model. Using the more traditional advantages of Xenopus, others are providing important lineage data on the origin of the cells that make up the tissues of the regenerate. Finally, transcriptome analyses of regenerating tissues seek to identify the genes and cellular processes that enable successful regeneration.

BackgroundDedifferentiation occurs naturally in mature cell types during epimorphic regeneration in fish and some amphibians. Dedifferentiation also occurs in the induction of pluripotent stem cells when a set of transcription factors (Oct4, Sox2, Klf4 and c-Myc) is over expressed in mature cell types.ResultsWe hypothesised that there are parallels between dedifferentiation or reprogramming of somatic cells to induced pluripotent stem cells and the natural process of dedifferentiation during epimorphic regeneration. We analysed expression levels of the most commonly used pluripotency associated factors in regenerating and non-regenerating tissue and compared them with levels in a pluripotent reference cell. We found that some of the pluripotency associated factors (oct4/pou5f1, sox2, c-myc, klf4, tert, sall4, zic3, dppa2/4 and fut1, a homologue of ssea1) were expressed before and during regeneration and that at least two of these factors (oct4, sox2) were also required for normal fin regeneration in the zebrafish. However these factors were not upregulated during regeneration as would be expected if blastema cells acquired pluripotency.ConclusionsBy comparing cells from the regeneration blastema with embryonic pluripotent reference cells we found that induced pluripotent stem and blastema cells do not share pluripotency. However, during blastema formation some of the key reprogramming factors are both expressed and are also required for regeneration to take place. We therefore propose a link between partially reprogrammed induced pluripotent stem cells and the half way state of blastema cells and suggest that a common mechanism might be regulating these two processes.

We have employed transgenic methods combined with embryonic grafting to analyse the mechanisms of regeneration in Xenopus tadpoles. The Xenopus tadpole tail contains a spinal cord, notochord and segmented muscles, and all tissues are replaced when the tail regenerates after amputation. We show that there is a refractory period of very low regenerative ability in the early tadpole stage. Tracing of cell lineage with the use of single tissue transgenic grafts labelled with green fluorescent protein (GFP) shows that there is no de-differentiation and no metaplasia during regeneration. The spinal cord, notochord and muscle all regenerate from the corresponding tissue in the stump; in the case of the muscle the satellite cells provide the material for regeneration. By using constitutive or dominant negative gene products, induced under the control of a heat shock promoter, we show that the bone morphogenetic protein (BMP) and Notch signalling pathways are both essential for regeneration. BMP is upstream of Notch and has an independent effect on regeneration of muscle. The Xenopus limb bud will regenerate completely at the early stages but regenerative ability falls during digit differentiation. We have developed a procedure for making tadpoles in which one hindlimb is transgenic and the remainder wild-type. This has been used to introduce various gene products expected to prolong the period of regenerative capacity, but none has so far been successful.