REGULATION OF INITIATION OF DNA SYNTHESIS IN CHINESE HAMSTER CELLS

Abstract

Suspension cultures of Chinese hamster cells (line CHO), which had stopped dividing and were arrested in G1 following growth to high cell concentrations in F-10 medium, could be induced to reinitiate DNA synthesis and to divide in synchrony upon addition of the appropriate amounts of isoleucine and glutamine. Both amino acids were required to initiate resumption of cell-cycle traverse. Deficiencies in other amino acids contained in F-10 medium did not result in accumulation of cells in G1, indicating a specific action produced by limiting quantities of isoleucine and glutamine. In the presence of sufficient glutamine, approximately 2 x 10-6 M isoleucine was required for all cells to initiate DNA synthesis in a population initially containing 1.5 x 105 cells/ml. Under similar conditions, about 4 x 10-6 M isoleucine was required for all G1-arrested cells to progress through cell division. In contrast, 1 x 10-4 M glutamine was necessary for maximum initiation of DNA synthesis in G1 cells, along with sufficient isoleucine. A technique for rapid production of G1-arrested cells is described in which cells from an exponentially growing population placed in F-10 medium deficient in both isoleucine and glutamine or isoleucine alone accumulated in G1 after 30 hr.