Emetine is an antibiotic which inhibits the activity of the 40-S subunits of eukaryotic ribosomes. A number of emetine-resistant mutants of Chinese hamster cells have been isolated. We have studied the ribosomes isolated from strains carrying either of two alleles of an emetine resistance locus, one of which leads to an alteration in the electrophoretic mobility of ribosomal protein S14. Both mutations lead to a loss of specific proteins, including S14, from 40-S subunits isolated in 0.5 M KCl, an effect which is enhanced at elevated temperature. 40-S subunits from wild-type cells retain their proteins even at substantially higher concentrations of KCl. Interestingly, the two mutant alleles lead to the loss of slightly different sets of proteins from the 40-S subunits. These results suggest that protein S14 plays an important role in the structure of the ribosome and raise the possibility that resistance to emetine is accomplished through an alteration in a complex emetine-binding site rather than in a single emetine-binding protein.