The class I surface Ag recognized during tissue graft rejection are composed of a 40- to 45-kDa H chain and a 12-kDa L chain, beta 2-microglobulin (beta 2m). Regulation of MHC gene expression during early development is thought to play an essential role in maternal tolerance of the fetal allograft. Here we used in situ hybridization techniques to characterize the temporal and spatial pattern of expression of MHC class I mRNA in the developing mouse embryo. MHC class I transcripts were initially detected at day 9.5 postcoitus in the primary and secondary trophoblast giant cell populations. At this stage, none of the remaining fetal components of the developing placenta expresses MHC class I mRNA. By contrast, the outer zone of trophoblast giant cells is the only trophectoderm-derived tissue in the developing placenta that does not express detectable levels of beta 2m mRNA. These findings indicate that the onset of MHC class I and beta 2m gene expression in early postimplantation stage embryos is not coordinately regulated. In addition, we made use of pluripotent embryonic stem (ES) cells to study developmental regulation of MHC class I and beta 2m gene expression in vitro. ES cells can be induced to form endoderm-like cells by the technique of suspension culture. In contrast to retinoic acid-treated F9 cells, the endoderm derived from ES cells expressed high levels of beta 2m mRNA, but no detectable MHC class I transcripts. These findings are consistent with results of our in situ hybridization experiments showing high levels of expression of beta 2m mRNA, and extremely weak expression of MHC class I transcripts in derivatives of the visceral endoderm in vivo.