The cDNA coding for the yellow fever virus (YFV) envelope protein (E) was inserted into an SV40 vector under the control of the late promoter in place of the VP1 gene. The recombinant virus expressed a 52-Kd polypeptide which was detected by immunoprecipitation with a monoclonal antibody raised against the E protein. Surprisingly, this protein was visualized in the nucleus of the infected cells. The possible presence of a sequence involved in nuclear migration of the E protein and naturally ignored during virus infection is discussed. The sequence coding for the mature E protein is directly preceded in the YFV genome by a sequence of 45 nucleotides coding for a 15-amino-acid long hydrophobic oligopeptide. The sequence of this oligopeptide was inserted into the SV40 recombinant virus between the ATG codon and the first codon for the E protein. The E protein expressed by this new SV40 recombinant virus was found to be localized in the cytoplasm of the infected cells in association with intracellular membranes. These results strongly suggest that the 15-amino-acid long hydrophobic region naturally plays a role as a signal peptide for the E protein.