NUCLEOCYTOPLASMIC RATIO REQUIREMENTS FOR THE INITIATION OF DNA REPLICATION AND FISSION IN TETRAHYMENA

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Abstract
Hydroxyurea [HU] (10 mM) arrests the exponential growth of Tetrahymena by blocking DNA replication during S-phase. After removal of the HU, a long recovery period occurs during which active DNA synthesis takes place. 3H-TdR [thymine deoxyribose] 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 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 h exposure to HU, N remains essentially the same whereas C increases. When the HU is removed, N increases by 4.times. and C continues to increase until just prior to recovery division when it also reaches a value 4.times. 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.