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shhxenopus hindlimb digit 

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Experiment details for shh

Gremlin1 induces anterior-posterior limb bifurcations in developing xenopus limbs but does not enhance limb regeneration.

Gremlin1 induces anterior-posterior limb bifurcations in developing Xenopus limbs but does not enhance limb regeneration.

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Gene Clone Species Stages Anatomy
shh.L laevis NF stage 55 hindlimb digit 5

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  Fig. 1. Grem1 expression, relative to shh and fgf8 supports the two phase limb reciprocal signalling model of Verheyden and Sun (2008) in Xenopus. A–W in situ hybridisation (dark purple staining) of developing (A–L) and regenerating Xenopus laevis limb buds. Note that the posterior limb bud is uppermost as this reflects the posture in which the embryo develops; the tadpole itself is anterior to the left and dorsal uppermost, lying on its side. This orients the limb proximal to the left. Some embryos were pigmented and show black melanophores. A–G) Shh in developing limbs is localised to the ZPA, the posterior distal mesenchyme, as previously shown. Expression appeared strongest at stage 52 and then declined up to stage 55, where it was found in digit V mesenchyme. H–L) Grem1 expression was first seen in distal mesenchyme at stage 50 (H), before becoming cleared from the mesenchyme directly under the AER by stage 51 (I). This trend continued at stage 52 (J) before gradual loss of expression in the autopod so that no transcripts were detectable by stage 54 (K, L). Black arrowheads in C and J indicate amputation site used for regenerating limb experiments and roman numerals indicate digit identity. M–Q) Expression of shh in regenerating limbs amputated at stage 52, midway through the limb bud. Shh was first detected at 2 dpa and was always localised to the posterior distal mesenchyme under the AEC (N–Q). By 6 dpa it was starting to reduce, as the autopod re-differentiated. R–W) Grem1 expression in regenerating limb buds appeared at 1 dpa and from 2 to 4 dpa was localised in distal anterior mesenchyme, reciprocal to shh (R–U). Expression cleared from the autopod at 5–6 dpa (V, W). X) Model of Verheyden and Sun (2008) based on genetic manipulations in mice. Y) Xenopus model showing alignment with the mouse model, FGF data from Wang and Beck (2014) for fgf8. Note limb drawings have been “inverted” to put anterior uppermost as is the convention for amniotes, and black arrowheads indicate amputation level. Grem1 is initially directly in contact with the AER and a positive feedback loop is established between Grem1 and Shh in the mesenchyme and the AER FGFs. This is phase I and corresponds to stage 50 of Xenopus limb development. At stage 51, grem1 is cleared from the distal mesenchyme because fgf levels in the AER rise above the threshold required to switch on an inhibitory loop (phase II). Shh levels continue to rise but grem1 is now distant from the AER Fgfs and may fail to maintain expression (stage 52). This leads to termination of the loop starting at stage 53 and loss of first grem1 and then shh and fgf8 from the autopod. Regenerating limbs are similar although since grem1 is not cleared from the distal mesenchyme until 5 dpa, phase I may last longer, enabling extra growth required to regenerate lost structures. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)