Differential display of genes expressed at the midbrain-hindbrain junctionidentifies sprouty2: An FGF8-inducible member of a family of intracellularFGF antagonists

specification and polarization of the midbrain and anterior hindbrain invol ve planar signals originating from the isthmus. Current evidence suggests t hat FGF8, expressed at the isthmus, provides this patterning influence. In this study, we have sought to identify novel genes which are involved in th e process by which regional identity is imparted to midbrain and anterior h indbrain (rhombomere 1). An enhanced differential display reverse transcrip tion method was used to clone cDNAs derived from transcripts expressed spec ifically in either rhombomere 1 or midbrain during the period of isthmic pa tterning activity. This gene expression screen identified 28 differentially expressed cDNAs. A clone upregulated in cDNA derived from rhombomere 1 tis sue showed a 91% identity at the nucleotide level to the putative human rec eptor tyrosine kinase antagonist: sprouty2. In situ hybridization on whole chick embryos showed chick sprouty2 to be expressed initially within the is thmus and rhombomere 1, spatially and temporally coincident with Fgf8 expre ssion. However, at later stages this domain was more extensive than that of Fgf8. Introduction of ligand-coated beads into either midbrain or hindbrai n region revealed that sprouty2 could be rapidly induced by FGF8. These dat a suggest that sprouty2 participates in a negative feedback regulatory loop to modulate the patterning activity of FGF8 at the isthmus.