M. Hola et al., INITIATION OF DNA-SYNTHESIS BY NUCLEI FROM SCRAPE-RUPTURED QUIESCENT MAMMALIAN-CELLS IN HIGH-SPEED SUPERNATANTS OF XENOPUS EGG EXTRACTS, Journal of Cell Science, 107, 1994, pp. 3045-3053
Demembranated sperm heads, detergent-isolated somatic nuclei and even
naked DNA are efficiently replicated in cytoplasmic extracts of activa
ted amphibian eggs, but only after nuclear assembly and the formation
of an intact nuclear envelope. DNA synthesis has not previously been s
hown to be initiated in high-speed (200,000 g) supernatants of egg cyt
oplasm because they are depleted of the vesicular material required to
support nuclear envelope formation. Here we show that mammalian nucle
i prepared by scrape-rupture are able to initiate DNA replication in s
uch high-speed supernatants. These nuclei begin DNA synthesis asynchro
nously, This asynchrony cannot be attributed to differences in the tim
e taken for nuclear assembly, Instead, we suggest that the asynchrony
reflects intrinsic differences between nuclei and that these differenc
es are a major cause of cell cycle variability. Our demonstration of i
nitiation in high-speed supernatants now enables the initiation of euk
aryotic DNA synthesis to be studied independently of nuclear assembly.