Differential replication capacities of G1 and S-phase extracts from sea urchin eggs

Abstract

Sea urchin eggs are arrested in G1 of the first mitotic cell cycle. Fertilization triggers release from G1 arrest and the onset of DNA synthesis about 20 minutes later, even when protein synthesis is blocked. Here we describe extracts from eggs and S-phase embryos that reproduce this stage-specific pattern of DNA synthesis. Fertilized egg extracts formed nuclear membranes around decondensed Xenopus sperm chromatin whereas unfertilized egg extracts did not. Aphidicolin-sensitive deoxynucleotide incorporation was high in extracts of fertilized S-phase eggs and low in those of unfertilized eggs. In contrast, single-stranded DNA templates directed high rates of incorporation in both unfertilized and fertilized egg extracts, suggesting that the stage-specific activities in nuclear DNA synthesis is restricted to initiation on double-stranded DNA. Mixing experiments showed that unfertilized eggs do not contain a dominant inhibitor of replication, nor does fertilization induce the appearance of a soluble, dominant activator.