Phosphorylation of ribosome-associated proteins during the mammalian cell cycle. Unique phosphorylation of a specific protein during mitosis.

Chinese hamster ovary cells in monolayer culture were incubated with [32P] phosphate. Ribosome-associated proteins, including both structural proteins and those tightly bound to washed, centrifuged ribosomes, were isolated and separated by electrophoresis on sodium dodecyl sulfate-polyacrylamide gels. The electrophoretic pattern showed five major regions or peaks of 32P radioactivity which represented phosphorylated ribosome-associated proteins with molecular weights of 17 500, 23 000, 30 000, 38 000, and 57 000. When asynchronous cells were pulse-labeled with [32P] phosphate, the predominant peak of 32P radioactivity was associated with the protein of 38 000 daltons. Similar results were obtained with cells synchronized in the G1, S, or G2 phase of the mammalian cell cycle. Conversely, proteins isolated from the ribosomes of mitotic cells, collected and labeled with [32P] phosphate in the presence of colcemid, showed a new and predominant peak of 32P radioactivity migrating with a protein of 45 000 daltons. When cells labeled in mitosis were allowed to progress into G1 phase, this peak of 32P radioactivity rapidly disappeared from the electrophoretic pattern. These results suggest that a specific protein associated with the ribosomes was phosphorylated uniquely during the mitotic phase of the cell cycle.