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Presenting Author(s) Eva Tiecke
Abstract Title A dual role for PKA in chick limb development.
Full author List Eva Tiecke, Juan-Jose Sanz-Ezquerro, Anne Warner and Cheryll Tickle
Text of abstract PKA is a cAMP-dependent kinase that, in Drosophila, has been shown to have a dual role in Hh signalling, through the processing of Ci. Recent work on limb development in mice has made it clear that whether Gli3, a vertebrate homolgue of Ci, is processed or not important for digit formation. We studied the role of PKA activity in the developing chick limb bud by altering levels of PKA. We added forskolin, a drug that increases PKA and has previously been reported to have a negative effect on Shh signalling, to cultures of cells from anterior quail limb buds and grafted them back to the anterior margin of chick limb buds. We also grafted paper soaked in forskolin to the anterior limb bud and infected chick limb buds with dominant negative and constitutively active PKA viruses. Forskolin treatment had little effect on the duplicating activity of posterior limb bud cultures but unexpectedly anterior limb bud cultures soaked in forskolin gave extra digits. Both host and graft cells make up part of the duplicated digit and forskolin treatment also enhanced gap junctional communication. Expression of downstream targets of Shh signalling gli1, ptc, hoxa13 and gremlin show that the Shh signalling pathway is activated by forskolin in cultured anterior cells, even though no Shh can be detected and we also found that levels of Gli3A and Gli3R are increased. When presumptive limb buds were infected with virus, we generated extra digits anteriorly both with activated and dominant negative PKA. In normal limb buds, PKA activity is high in the posterior and low in the anterior of the limb bud. Taken together, these results suggest a dual role for PKA in limb development not only in repressing Shh signalling but also in activating Shh signalling. PKA activity in the limb may be mediated by cAMP which also regulates gap junctional communication.
Which session is your work most relevant to: Limb initiation, Limb patterning, Limb regeneration, Tissue differentiation, Vascularisation & innervation, Human limb abnormalities, Limb evolution