Pathogenic Yersinia species have a specialized secretion system (type III) to target cytotoxic Yap proteins during infection. The signals of YopE and YopN sufficient for the secretion of translational reporter fusions were mapped to the first 15 codons, No common amino acid or peptide sequence could be identified among the secretion signals. Systematic mutagenesis of the secretion signal yielded mutants defective in Yop translation; however, no point mutants could be identified that specifically abolished secretion. Frameshift mutations that completely altered the peptide sequences of these signals also failed to prevent secretion. Thus, the signal that leads to the type III secretion of Yop proteins appears to be encoded in their messenger RNA rather than the peptide sequence.