We have developed a sensitized screen to identify genes involved in gene silencing, using random N-ethyl-N-nitrosourea mutagenesis on mice carrying a variegating GFP transgene. The dominant screen has produced six mutant lines, including both suppressors and enhancers of variegation. All are semidominant and five of the six are homozygous embryonic lethal. In one case, the homozygous lethality depends on sex: homozygous females die at midgestation and display abnormal DNA methylation of the X chromosome, whereas homozygous males are viable. Linkage analysis reveals that the mutations map to unique chromosomal locations. We have studied the effect of five of the mutations on expression of an endogenous allele known to be sensitive to epigenetic state, agouti viable yellow. In all cases, there is an effect on penetrance, and in most cases, parent of origin and sex-specific effects are detected. This screen has identified genes that are involved in epigenetic reprogramming of the genome, and the behavior of the mutant lines suggests a common mechanism between X inactivation and transgene and retrotransposon silencing. Our findings raise the possibility that the presence or absence of the X chromosome in mammals affects the establishment of the epigenetic state at autosomal loci by acting as a sink for proteins involved in gene silencing. The study demonstrates the power of sensitized screens in the mouse not only for the discovery of novel genes involved in a particular process but also for the elucidation of the biology of that process.