Is the Nuclear Membrane Involved in DNA Replication?

Abstract

In 1963 Jacob, Brenner and Cuzin proposed the replicon concept for bacterial DNA and on the basis of some later modifications suggested that the replicating point was attached to the bacterial cell membrane. Since basic mechanisms in prokaryotes have a certain probability of being similar to basic mechanisms in eukaryotes, I was interested in, examining the question of whether membrane attachment played a role in DNA replication in mammalian cells. We were primarily interested in whether DNA replication might be initiated at the nuclear membrane since a few autoradiographic studies had already suggested that replication could take place throughout the nucleus, implying that in mammalian cells the replication point did not always remain at the membrane. These previous studies, however, were mostly with 30 minute pulse labels. To examine this question human amnion cells were synchronized first with excess thymidine for 24 hours. The excess thymidine was then washed out and the cells allowed to pass through the S period for 10 hours and then further synchronized by exposure to amethopterin for 14 hours. The cultures were then exposed to tritiated thymidine for 10 minutes and after appropriate fixation electron microscope autoradiography performed.