Ectopic Wnt signal determines the eyeless phenotype of zebrafish masterblind mutant

masterblind (mbl) is a zebrafish mutation characterised by the absence or r eduction in size of the telencephalon, optic vesicles and olfactory placode s. We show that inhibition of Gsk3 beta in zebrafish embryos either by over expression of dominant negative dn gsk3 beta mRNA or by lithium treatment a fter the midblastula transition phenocopies mbl. The loss of anterior neura l tissue in mbl and lithium-treated embryos is preceded by posteriorization of presumptive anterior neuroectoderm during gastrulation, which is eviden t from the anterior shift of marker genes Otx2 and Wnt1. Heterozygous mbl e mbryos showed increased sensitivity to inhibition of GSK3 beta by lithium o r dn Xgsk3 beta that led to the loss of eyes. Overexpression of gsk3 beta m RNA rescued eyes and the wild-type fgf8 expression of homozygous mbl embryo s. emx1 that delineates the telencephalon is expanded and shifted ventroant eriorly in mbl embryos. In contrast to fgf8, the emx1 expression domain was not restored upon overexpression of gsk3 beta mRNA. These experiments plac e mbl as an antagonist of the Wnt pathway in parallel or upstream of the co mplex consisting of Axin, A-PC and Gsk3 beta that binds and phosphorylates beta -catenin, thereby destabilising it mbl maps on LG 3 close to a candida te gene axin1. In mbl we detected a point mutation in the conserved minimal Gsk3 beta -binding domain of axin1 leading to a leucine to glutamine subst itution at position 399. Overexpression of wild-type axin1 mRNA rescued mbl completely, demonstrating that mutant axin1 is responsible for the mutant phenotype. Overexpression of mutant L399Q axin1 in wild-type embryos result ed in a dose-dependent dominant negative activity as demonstrated by the lo ss of telencephalon and eyes. We suggest that the function of Axin1/Mbl pro tein is to antagonise the Wnt signal and in doing so to establish and maint ain the most anterior CNS. Our findings provide new insights into the mecha nisms by which the Wnt pathway generates anteroposterior polarity of the ne ural plate.