Tumor-suppressor p53 regulates cell cycle, differentiation and apoptosis. The effect of all-trans-retinoic acid (RA) on p53, other protooncogenes and downstream signaling molecules were investigated during murine development in the following studies. Temporal modulation of p53 in murine embryonic stem cells (ES) was investigated after exposure to 1 M RA on day 8, 9 and 10 of differentiation. ES cells are pluripotent cells derived from the inner cell mass of blastocysts and can spontaneously differentiate into cells of various lineages. ES cells also respond to various internal and external signals of proliferation and differentiation and can thus mimic the in vivo differentiation process. Undifferentiated ES cells expressed high levels of p53 mRNA and protein that declined as differentiation proceeded. Increased apoptosis was observed after RA treatment. In RA-treated cells, a transient increase in the expression of p53 and c-myc mRNA and protein was correlated with neural differentiation. Increased expression of Max and Mad genes along with elevated c-myc in RA-treated ES cells suggested that RA could modulate Myc family for effective signaling. Increased expression of caspase 3, caspase 6, Bax, and Bad was seen during RAmediated ES cell differentiation. Increased expression of tumor necrosis factor , macrophage migration inhibitory factor and IL-18 were also noted in RA-treated cells. To investigate the effects of RA during murine organogenesis, assessment of p53 and Myc family was undertaken in Swiss Webster mice fetuses after maternal treatment with 100 mg/kg of oral RA. In utero RA-exposure resulted in decreased expression of p53 mRNA and protein in a temporal and spatial manner in the fetuses. Temporal modulation of Myc family and other pro-apoptotic genes were also noted after in utero RA-exposure. Increased apoptosis was noted in RA-treated fetuses at 24-48 h, which declined by 72 h. These studies demonstrated that RA can temporally modulate the expression of p53 and the Myc gene family during in vitro ES cell differentiation and in vivo murine development. A basic understanding of embryonic development requires an appreciation of the complexity involved in proto-oncogene and tumor-suppressor signaling that, in turn, can facilitate understanding the loss of orderly control during oncogenic transformation.