Hydroxyurea (10 mM) arrests the exponential growth of Tetrahymena by blocking DNA replication during S‐phase. After removal of the hydroxyurea (HU), they have a long recovery period during which they are active in DNA synthesis. 3H‐TdR uptake showed that on completion of the recovery period, the cells divide (recovery division) and enter a cell cycle which lacks G1. The frequency, size and DNA content of the extranuclear chromatin bodies (ECB) formed at this division are all markedly increased (2–4) over the corresponding values obtained from exponential growth phase controls. Microspectrophotometric analysis of macronuclear DNA content (N) coupled with the cytoplasmic dry mass (C) values suggest that specific N to C ratios (N/C) are required for the initiation of DNA replication and fission: during a normal (exponential growth) cell cycle, both N and C double, but asynchronously, so that the N/C of both post‐fission‐daughter cells and pre‐fission cells is identical (standardized to N/C = 1) but late G1 cells have a low N/C. During a 10 hr exposure to HU, the N remains essentially the same whereas the C increases. When the HU is removed, the N increases by 4× and the C continues to increase until just prior to recovery division when it also reaches a value 4× that of the original daughter cells. Thus, the N/C = 1 is re‐established. The enlarged ECB formed during recovery division may function to lower the N/C in the daughter cells, which in turn may in some way stimulate immediate DNA replication, thus eliminating G1. The elimination of G1 (and shortening in a few subsequent cell cycles) allows less time for cytoplasmic growth and results in the return of the cells to the generation time and the N and C values observed prior to the HU treatment.
Observation of division of individual cells in microdrops, plus autoradiographic studies using tritiated thymidine and standard cell cycle analysis techniques, reveal that hydroxyurea (10 mM) reversibly arrests the normal progression of exponentially growing Tetrahymena pyriformis through the initial 92 % of S-phase while not affecting cells in the terminal 8 % and in G, and division. Thus the fraction of the population of cells that is in G2 can be approximately determined by the fractiofi of the population able to divide in the presence of hydroxyurea. This fraction can be related to the approximate duration of G2 by calculations which compensate for the age gradient.
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