THE PREGASTRULA ESTABLISHMENT OF GENE-EXPRESSION PATTERN IN XENOPUS EMBRYOS - REQUIREMENTS FOR LOCAL CELL-INTERACTIONS AND FOR PROTEIN-SYNTHESIS

Although it has been proposed that mesoderm forms in the marginal zone of the amphibian embryo through inductive signaling from vegetal pole cells, the details of this process remain to be clarified. To determi ne when marginal zone cells become committed to mesodermal fates, cell contact and protein synthesis requirements for early transcriptional responses were analyzed in Xenopus blastulae. Marginal zone explants w ere isolated from embryos at different stages and either were cultured untreated, or were dissociated in a medium lacking calcium and magnes ium ions, or cultured in the presence of cycloheximide. Whereas many m esoderm-specific transcripts are efficiently induced by activin in dis sociated animal pole cells, the same markers were not activated in dis sociated marginal zone cells, which were isolated from mid or even lat e blastulae and cultured in the absence of exogenous inducers. These o bservations suggest that early specification of mesodermal fates requi res cell-cell interactions within the marginal zone and that the margi nal zone cells are not committed to express early mesodermal markers u ntil the late blastula stage. Specification of mesodermal fates was al so assessed by the ability of marginal zone cells to express early mes odermal markers in the absence of protein synthesis. Induction of Xlim 1, 1A11, and, partially, Xbrachyury transcripts in the marginal zone w as blocked by cycloheximide treatment through late blastula stages, wh ereas Goosecoid and Xwnt8 mRNAs were expressed in the absence of prote in synthesis, indicating that these sets of markers are activated in v ivo through different pathways. These observations demonstrate that de termination of mesoderm in the marginal zone is a multistep process wh ich occurs during late blastula stages and depends on cell-cell contac t and protein synthesis. (C) 1994 Academic Press, Inc.