USE OF AN OOCYTE EXPRESSION ASSAY TO RECONSTITUTE INDUCTIVE SIGNALING

We have developed a paracrine signaling assay capable of mimicking ind uctive events in the early vertebrate embryo, RNA encoding one or more secreted proteins is microinjected into a Xenopus laevis oocyte, Afte r a brief incubation to allow translation, a piece of embryonic tissue competent to respond to the signaling protein is grafted onto the ooc yte, The secreted protein's effect on the grafted explant is then scor ed by assaying expression of tissue-specific markers, Explants of ecto dermal tissue from blastula or gastrula stage embryos were grafted ont o oocytes that had been injected with RNA encoding activin or noggin, We found that the paracrine assay faithfully reconstitutes mesoderm in duction by activin and neural induction by noggin, Blastula-stage expl ants grafted onto activin-expressing oocytes expressed the mesodermal marker genes brachyury, goosecoid, and muscle actin, Gastrula-stage ex plants grafted onto noggin-expressing oocytes expressed neural cell ad hesion molecule (NCAM) and formed cement gland, By injecting pools of RNA synthesized from a cDNA expression library into the oocyte, we als o used the assay to screen for secreted neural-inducing proteins, We a ssayed 20,000 independent transformants of a library constructed from LiCl-dorsalized Xenopus laevis embryos, and we identified two cDNAs th at induced neural tissue in ectodermal explants from gastrula-stage em bryos, Both cDNAs encode noggin, These results suggest that the paracr ine assay will be useful for the cloning of novel signaling proteins a s well as for the analysis of known factors.