FURTHER STUDIES ON THE PATTERN OF CHROMOSOME DUPLICATION IN CULTURED MAMMALIAN LEUCOCYTES.

Abstract

Autoradiographic studies on chromosome duplication in cultured mammalian cells (have revealed several very significant findings, including the detection of asyncronous DNA replication in homologous X-chromosomes. In cultured cells froin ,a female Chinese hamster, Taylor (1960) observed that the short arm of one of the X-chromosomes terminates duplication during the first half of the D N A synthetic period, but rhe long arm of the same X-chromosome and the entire portion of the other X-chromosome duplicates only in rhe later part of the D N A synthetic period. A somewhat similar type of out-ofphase D N A replication has also been observed very clearly in presumptive X-chromoson~es in cultured human cells (Ailorishima et nl., 1962; German, 1962; Rowley et nl., 1963). In cultured cells from normal females as well as in diploid polysomic-X cells only one X-chromosome conlpletes its replication along with the autosomes, while the remaining X-chromosome or chromosomes continue their replication after rhe rest of the conlpleillent has terminated duplication. Although the morphologic identification of the X-chron~osomes in Chinese hamster cells is conlparatively easy, it is not so for human X-chromosoines. Since no late-replicating chromosome in group 6-12 + X is seen in normal males as it is in normal females, such late-replicating chron~osomes are considered to be X-chromosomes. T,he sex cl~romarin lnasses found in interphase nuclei with lnore than one X-~hromosome are thought to be forined b) the heteropycnosis of the Xchromosomes (Ohno et nl., 1959), and it is further believed that the heteropycnotic-X-chronlosonles are the ones which complete their D N A replication later than all other chromoson~es in the colnplement (Grumbach et nl., 1963; Atkins et nl., 1962). In general, the number of late-labelling X-chroinosornes observed has been equal to the maximum number of sex chromatin masses present in inter~hase nuclei and one less than the number of X-chroi~losomes in the L diploid complement. Based on the evidence from mouse genetics, Lyon (1961) has postulated that the heteropycnotic X-chroil~osome found in normal mammalian females is genetically inactive, and this ll~ypothesis is supported by cytological observations in cells with an X-autosome translocation in mice (Ohno and Cattanach, 1962) as well as by less direct evidence of glucose-6 phosplhate dehydrogenase activity in inan (Beutler et al., 1962). Pooling all the available information, it appears now that the X-chromosome which forms the sex chromatin mass in interphase nuclei, the heteropycnotic-X, the latereplicating-X and the inactive X are indeed the different manifestations of the same X-chromosome. In mammalian chromosome comuleinents. the X-chromosomes are not rhe I , only ones which show a definite Dattern of D N A re~lication. The autosomes I I and the Y-chromosome also appear to follow a ,definite time and morphologic sequence, and even homologous autosomes may show characteristically differ-