GSK3β controls the timing and pattern of the fifth spiral cleavage at the 2–4 cell stage in Lymnaea stagnalis

Abstract

Establishment of the body plan of multicellular organisms by the primary body axis determination and cell-fate specification is a key issue in biology. We have examined the mRNA localization of three Wnt pathway components gsk3β, β-catenin, and disheveled and investigated the effects of four selective inhibitors of these proteins on the early developmental stages of the spiral cleavage embryo of the fresh water snail Lymnaea (L.) stagnalis. mRNAs for gsk3β and β-catenin were distributed uniformly throughout the embryo during development whereas disheveled mRNA showed specific localization with intra- and inter-blastomere differences in concentration along the A-V axis during spiral cleavages. Remarkably, through inhibitor studies, we identified a short sensitive period from the 2- to 4-cell stage in which GSK3β inhibition by the highly specific 1-azakenpaullone (AZ) and by LiCl induced a subsequent dramatic developmental delay and alteration of the cleavage patterns of blastomeres at the fifth cleavage (16- to 24-cell stage) resulting in exogastrulation and other abnormalities in later stages. Inhibition of β-Catenin or Disheveled had no effect. Our inhibitor experiments establish a novel role for GSK3β in the developmental timing and orientated cell division of the snail embryo. Further work will be needed to identify the downstream targets of the kinase.