Mechanisms of cell cycle regulation in embryos of normal and unusual size - PROJECT SUMMARY Mammalian development is a remarkably robust process that employs self-organization and corrective processes to coordinate individual cell behaviors for proper development. One example is the embryo’s capacity for cell number regulation. In mouse embryos in which cell number is experimentally doubled (2x), cell number correction was shown to occur shortly after the embryo implants into the uterine wall and was suggested to involve lengthening of cell cycles. This early postimplantation period is known to be a time of dramatic transitions where a slowly proliferating clump of cells that makes up the embryonic portion of the conceptus - the epiblast (EPI) - transforms into a rapidly dividing polarized epithelium that demonstrates interkinetic nuclear migrations (IKNM) in conjunction with the cell cycle. Previous studies analyzing fixed 2x embryo samples have described abnormalities in polarization and epithelial organization of these embryos, however, how these morphogenetic differences influence cell cycle dynamics remains unknown. The proposed study will utilize ex vivo live imaging of early postimplantation embryos and stem cell models with a novel fluorescent cell cycle reporter to investigate the details of this cell cycle slowing as it relates to cell number regulation, determine how cell cycle regulation is rooted in morphogenetic abnormalities and probe the molecular link between the two processes. Aim 1 will investigate the requirements for proper morphogenesis in the initiation and maintenance of rapid cell cycles and use an embryonic stem cell model to precisely test the mechanistic links between the two. Aim 2 will investigate the polarization defects in 2x embryos, precisely quantify the cell cycle remodeling thought to underlie size regulation and attempt to overcome slowed cell cycles by artificially accelerating morphogenesis. Ultimately, these studies will be the first to describe phase-resolved dynamics of the cell cycle at this developmental period and increase our understanding of the embryo-intrinsic regulatory mechanisms that govern mammalian development. My training plan is precisely designed to develop the technical and professional skills that will support me in becoming an interdisciplinary independent researcher. Princeton University and the Posfai lab specifically possess the advanced facilities and collaborative, intellectual environment ideal for achieving this goal. My developmental biology training, overseen by my supportive sponsor and co-sponsor and enriched by the expertise of collaborators such as Dr. Stanislav Shvartsman and Dr. Daniel Cohen, will increase my skillset in live imaging, embryology, computational approaches, and quantitative data analysis. Combined with my access to professional training including frequent presentations in colloquia, workshops covering manuscript writing and computational skills, exposure to science communication, and my outreach work, this plan will ensure the success of my training.
