During different periods of mammalian development, global changes in gene expression occur. Developmental changes in global gene expression have been modeled as a restrictive process. To test the restriction model of global changes in gene expression, we have used embryonic stem (ES) cells as a model system for the early mammalian embryo. ES cells are pluripotent cells that can contribute to all cellular lineages of the developing mammalian fetus and are derived from early embryonic cells. Using this model system, we have studied a new class of RNAs called microRNAs that have been identified and shown to play a role in the direct regulation of messenger RNAs. Here we report the expression signature for 248 microRNAs in 13 independent murine ES cells, embryoid bodies, and somatic tissues. The expression profile for 248 mouse microRNAs was determined for embryonic stem cells, embryoid bodies, mouse embryos, mature heart, lung, liver, kidney, and brain. Characteristic microRNA expression signatures were observed for each evaluated sample. When the characteristic microRNA signatures for developmentally ordered samples were compared, immature samples exhibited a less complex microRNA transcript profile than did mature samples. Our data support a progressive model of microRNA gene expression. Based on the progressive increase in complexity of micro- RNA expression, we hypothesize that the mammalian developmental program requires a temporal coupling of expression between microRNAs and messenger RNAs to enable the developmental potential observed in mammalian ontogeny.