Asynchrony of DNA synthesis

Moorhead, P. S., and V. Defendi. 1963. Asynchrony of DNA synthesis in chromosomes of human diploid cells. J. Cell Biol., 16 202-208. 

Asynchrony of DNA Synthesis During the Period of Heat Shocks. 

A very clear picture of asynchrony and local heterogeneity of DNA synthesis in the chromosomes was obtained in studies of DNA synthesis in the polytene chromosomes of insect salivary glands. We shall describe the characteristics of synthesis of DNA, RNA, and proteins in these chromosomes in greater detail later in connection with our examination of chromosomal puffs. At this point, however, I shall mention the results of experiments (Plaut, 1963 Plaut and Nash, 1964) in which asynchrony of DNA synthesis in different parts of individual chromosomes was demonstrated very clearly. 

Stubblefield was able to study the incorporation of tritiated thymidine into the different karyomeres of the same cell. While each of the individual nuclei was able to synthesize DNA and to enter mitosis, the distribution of label in the interphase nuclei and in the chromosomes clearly demonstrated asynchrony of DNA synthesis in the different nuclei of a single cell. Thus, multinucleated interphase cells were found which contained label in only some of the karyomeres. Since all these karyomeres entered mitosis simultaneously, the (tetraploid) metaphases were characterized by the presence of both heavily labeled and completely unlabeled chromosomes (Note 10). This experiment shows that the coordinated replication of the chromosomes breaks down when the chromosomes are segregated into several nuclei, or, putting it the other way around, that the coordination of the cycles of individual chromosomes requires that they be located within a single nucleus. 

Asynchrony, that is to say different rates of DNA synthesis in different parts of the chromosomes, was investigated by incorporation of labeled thymidine into chromosomes of root cells of Tradescantia (Wimber, 1961) and Vida faba (Peacock, 1963 Woodard et al., 1961), into chromosomes of He La cells (Stubblefield and Mueller, 1962 Mueller and Kajiwara, 1966), into chromosomes of cultures of leukocytes and other cells of the Chinese hamster (Prescott and Bender, 1963), and in investigations on certain other organisms. 

The notion that the rate of DNA synthesis and of other events of the chromosomal cycle, and, hence, the duration of this cycle (rather than the time of initiation of DNA synthesis), is diflFerent in different karyomeres, which contain different numbers and kinds of chromosomes, is not farfetched. Moreover, it has two advantages (1) It resolves the apparent conflict between the observations of Stubblefield on the asynchrony of karyomeres and those on the synchrony of (complete) nuclei in bi- and multinucleate cells (Note 12) and (2) it makes it easier to visualize the coordination of the replication of all chromosomes in hybrids between cells differing in generation times. We shall therefore conclude that a single, basic rate and timetable of chromosomal changes is established only when all the chromosomes are enclosed in a single nuclear membrane. 

Stubblefield considers that the simplest explanation for the occurrence of such [asynchronous] cells was that they resulted from multinucleated cells with asynchronous DNA synthesis. The unlabeled chromosomes came from those nuclei that had finished DNA synthesis prior to the addition of the tritiated thymidine, and the labeled chromosomes came from nuclei that were still replicating DNA after the introduction of the isotopic precursor. He indicates however another possibility, namely that all nuclei may begin DNA synthesis together, but at differing rates, so that the asynchrony may be observed only near the end of the DNA synthesis period. ... 

This confiict can be resolved also by assuming that the signal initiating DNA synthesis originates in one or a few chromosomes of the complement and that it travels slowly through the cytoplasm. Such a hypothesis would also explain why a degree of asynchrony between chromosomes similar to that described by Stubblefield is not observed when all chromosomes are located within the same nucleus (rather than in different karyomeres). It becomes, in fact, very attractive when taken in conjunction with the possible role of the attachment of the chromosomes to the nuclear membrane (considered in Section IV, C). 

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