94 J. JIEHHEHT TAYLOR 



its phosphorylation. The mu-lt'osido inonoiiliosphatc hinds with the 

 enzyme that normally convei'ts deoxyui'idylate to thyniidylate and 

 bloeks synthesis. In the ahsnicc of thymidine tripliosijhate, DNA repli- 

 cation ceases (sec Chapter II. The root cells of Vicia faba are quite 

 sensitive to the inhibitor. At a coiicciiti-ation of 10 "il/ the rate of cell 

 division drops to a low Ic\t1 within (5 hours, l)ut tiie cells which arrive 

 at division have intact chromosomes. At higher concentrations {10''' M 

 and higher) the chromosomes show extensive damage. The damage 

 appears about 3 hours after immersion of roots in the inhibitor. Lesions, 

 which appear as non-staining gaps in the late prophase, metaphase, and 

 anaphase chromosomes, are the first visual signs of damage. Some of the 

 gaps are complete discontinuities of the chromatids and free fragments 

 are the results. Frequently, but not regularly, both chromatids are 

 affected at the same locus. Many of the last cells to arrive at division 

 show an extensive shattering of all or most of the chromosomes. The 

 lesions apparently occur in cells near the end of their DNA replication 

 cycle when the enzyme, thymidylate synthetase, is quickly blocked. Such 

 cells have no mechanism to prevent their progress through division even 

 though their DNA replication is not complete. The lesions can be pre- 

 vented by supplying exogenous thymidine along with the inhibitor. In 

 addition, the lesions can be cured by giving the cells thymidine at least 

 1 hour before anaphase, when they are presumably in late interphase 

 or early prophase. Bromodeoxyuridine can be substituted for thymidine 

 in the healing of these lesions. Evidently, DNA replication, which 

 requires all four nucleotides including thymidylate, is necessaiy for 

 healing the lesions. 



An interesting feature of the fragmentation is tlie absence of chroma- 

 tid exchanges or any aberrations that will produce bridges at anaphase. 

 This observation led to the idea that DNA replication might be required 

 for reunion of chromosomes broken by radiation. A test of the hypothesis 

 appeared possible because a concentration of 10" M stopped cell division 

 and presumably DNA replication without yielding visible chromosome 

 damage. By giving cells a dose of 2o r of X-irradiation 4 hours before 

 fixing, anaphase bridges were produced in 6.4% of the cells. If the cells 

 had been placed in 10"'^ ilf FUDR for 30 minutes before irradiation and 

 were grown in the same solution until fixed 4 hours later, only 1.1% 

 of the anaphases had bridges. By supplying tliyniidiiie or bromodeoxy- 

 uridine no-' M) to the irradiated cells treated with FUDR, the per- 

 centage of bridges could be restored to nearly the control level, 5.4 and 

 5.5%, respectively. From tliese experiments the hypothesis was projiosed 

 that reunion of broken chromosomes requires DNA replication. The 

 process was visualized as involving two steps. The first would be a very 



