Replication of a mammalian genome: the role of de novo protein biosynthesis during S phase

ts14 is a temperature-sensitive Chinese hamster lung cell mutant that ceases protein biosynthesis within a short time of transfer to nonpermissive temperature (Haralson and Roufa, 1975; Roufa and Haralson, 1975; Roufa and Reed, 1975). This mutant contains a revertible, presumably a point mutation that renders its 60S ribosomal subunit thermolabile (Haralson and Roufa, 1975). In this report, we describe the relationship between the conditional ability of ts14 to synthesize protein during S phase and the replication of its DNA.After transfer to nonpermissive temperature (39°C), where ts14 synthesizes protein at a rate approximately 20 fold less than wild-type cells, synchronous cultures of the mutant performed all the processes required for replication of their DNA. During prolonged incubations at nonpermissive temperature, S phase ts14 completed approximately one round of DNA replication semi-conservatively as judged by density-transfer experiments. Pulse-labeling experiments performed on S phase cells revealed that ts14 synthesized the intermediates of discontinuous DNA replication at nonpermissive and permissive temperatures at similar rates. In these tests, the mutant was not substantially different from wild-type at both culture temperatures. At the nonpermissive temperature, however, ts14 synthesized significantly less nuclear protein (that is, histone) than did wild-type cells, and the mutant's chromatin appeared deficient in histone by virtue of its increased sensitivity to nuclease.