Studying the DNA damage response in embryonic systems

Abstract

Maintenance and surveillance of genome integrity is crucial during the very early steps of embryonic development, since de novo mutations generated during this stage can be propagated in differentiated adult cells and may lead to predisposition to diseases including cancer. Surprisingly, early embryos are characterized by a relaxed control of genome integrity, reminiscent of that observed in cancer cells. How embryos manage to produce healthy adult individuals in such conditions remains still unclear. Here, we describe protocols and methods to study and analyze the DNA damage response and genome integrity in two embryonic experimental systems, early Xenopus laevis embryos and mouse embryonic stem cells. We describe methods to study gene functions in the DNA damage response by mRNA microinjection in Xenopus embryos generated by in vitro fertilization, mutagenesis and developmental regulation of the DNA damage response. We also describe methods to analyze the DNA damage response in mESCs, including synchronization experiments that allow studying the DNA damage response at different cell cycle stages. Analysis of genome integrity in these systems may also help to shed light on the molecular mechanisms that preserve genome integrity and become dysregulated in cancer cells.

Keywords: Chromatin; DNA damage; DNA replication; MMS; Mouse embryonic stem cells; Nucleus; UV; Xenopus.