Emr-2 is an emetine-resistant (Emr) Chinese hamster ovary cell mutant that contains a single electrophoretically altered ribosomal protein, a component of its 40S ribosomal subunit [Boersma, D., McGill, S., Mollenkamp, J. & Roufa, D. J. (1979) J. Biol. Chem. 254, in press]. This report describes a genetic experiment designed to test linkage between the genes that specify the altered ribosomal protein, S20*, and emetine resistance in Emr clones that segregated from cultures of Emr-2 cells hybridized with emetine-sensitive cells. The data described indicate that Emr and S20* phenotypes are due to mutations linked to the same chromosome in the Chinese hamster genome; most likely they are due to the same mutation. The data also confirm earlier speculations by others that the Emr locus in Chinese hamster cells is hemizygous. Several Chinese hamster cell mutants that express genetically altered ribosomal functions have been isolated in our laboratory. Within this collection are nine mutagen-induced clones whose ribosomes are resistant to emetine (Emr), an alkaloid drug inhibitor of protein biosynthesis (1). We used two-dimensional polyacrylamide gel electrophoresis to analyze 72 proteins extracted from Emr mutants' ribosomes. We observed that only one ribosomal protein prepared from mutant clone Emr-2 was altered in a manner detectable by our assay. The altered protein, S20*, a component of the 40S ribosomal subunit, differs from wild-type S20 protein in its electrophoretic migration at pH 5. S20 and S20* migrated with identical mobilities in buffer containing sodium dodecyl sulfate (i.e., appeared to possess the same molecular weight) (1). These observations were consistent with S20* having arisen by a point mutation. The identification of an altered 40S ribosomal subunit protein from an Emr clone agreed with other investigators' assignment of emetine resistance in Chinese hamster cells to the 40S ribosomal subunit (2).Three genetic models could account for the relationship between the altered ribosomal protein S20* and Emr-2's Emr phenotype: (i) Emetine resistance might be independent of the alteration in S20 protein.(ii) The mutation responsible for emetine resistance might affect a gene coding for a protein that modifies S20 protein. S20* thus might be the nonmodified, electrophoretically distinguishable precursor to normal S20. (iii) The structural gene that codes for S20 protein might contain a mutation responsible for both S20* and the mutant's resistance to emetine.The electrophoretic and genetic experiments described in this report were designed to investigate the relationship between S20* protein and the Emr phenotype of clone Emr-2. We fused a derivative of Emr-2 with emetine-sensitive (Ems) Chinese hamster cells and selected several approximately tetraploid hybrid clones. Spontaneous Emr mitotic segregants were isolated from hybrid clones, and proteins from the 40S ribosomal subunits of hybrid and segregant clones were analyzed by two-dimensional polyacrylamide gel electrophoresis. By these experi...

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