We investigated the requirements of the carboxyterminal sequence for surface expression of the influenza viral hemagglutinin (HA). Deletions in the cloned hemagglutinin gene were introduced at locations upstream from and spanning into the region that codes for the hydrophobic carboxyl terminus. Primate cells infected with recombinants of the deleted HA gene and an SV40 vector were negative for surface immunofluorescence and failed to adsorb erythrocytes. Polypeptide analysis showed that the mutant hemagglutinins lacking the normal hydrophobic carboxy-terminal sequences were secreted into the medium. These data provide evidence that these sequences of the influenza hemagglutinin are responsible for accumulation at the cell surface. During infection with each deletion mutant, a truncated HA polypeptide was found intracellularly. Both intracellular and extracellular HAs were glycosylated, since a third species representing the unglycosylated mutant hemagglutinin was detected in the presence of tunicamycin. Interestingly, the secreted and intracellular mutant HA polypeptides differ from the surface HA in their sensitivity to endoglycosidase H, indicating that an alteration of glycosylation has occurred.