Embryonic stem cells (ES cells) are pluripotent stem cells that can produce all cell types of an organism. ES cells proliferate rapidly and are believed to be subject to high intrinsic replication stress. A recent study in EMBO Reports by Kurashima and his team challenge this assumption by providing a detailed molecular study of replication dynamics in these cells.

Led by Dr. Tomomi Tsubouchi at the National Institute for Basic Biology (NIBB) in Japan, the research team investigated replication fork dynamics – such as replication fork speed, pause frequency and origin firing density – in different substages of S phase. Their findings reveal previously overlooked aspects of replication not only in ES cells but also in other non-pluripotent cells.

Dr. Kiminori Kurashima, the first author of the study, explains: “By fractionating cells at sub-stages of S phase and performing DNA fiber analyses on these sorted populations, we discovered that pluripotent mammalian stem cells maintain a slow fork rate and high active origin density throughout S phase, with minimal fork pauses.”

This is inconsistent with their finding that in non-pluripotent cells, fork velocity is slower at the beginning of S phase but increases thereafter. Fork pauses also occur in non-pluripotent cells, especially in early S phase, with checkpoint mechanisms likely activated to facilitate fork acceleration and reduce pauses.

The research also shows that after differentiation, mouse ES cells acquire replication properties similar to those of non-pluripotent cells. In addition, some features of DNA replication observed in mouse ES cells are identical to human iPS cells, suggesting that the slow replication fork with high origin density may be a hallmark of pluripotency. Surprisingly, forcing replication forks to accelerate resulted in miscoordination between the completion of genome replication and cell cycle progression.

Dr. Tsubouchi concludes: “We propose that slow replication forks are not the manifestation of replication obstructions, but rather an integral part of DNA replication in ES cells. Our study highlights the dynamic regulation of DNA replication and shows how different cell types employ different mechanisms.”