Molecular control of the oocyte-to-zygote transition
The challenge in fertilization is to transform two highly differentiated cells, egg and sperm, into a single totipotent cell - the zygote - with vastly different properties compared to its parental cells. To generate a zygote, the cell cycle machinery switches from meiosis to mitosis and chromatin is reorganized and reprogrammed. How these fascinating processes are regulated at the molecular level and how their deterioration impacts fertility are key questions we aim to address in our research.
The oocyte-to-zygote transition is one of the most dramatic cell conversions in biology. It refers to the female germ cell or oocyte, which undergoes two rounds of meiotic chromosome segregation and, following fertilization, is converted into a mitotically dividing embryo (Figure 1). We are addressing fundamental questions relating to the processes that ensure the inheritance of genomes from one generation to the next by combining germ cell and chromosome biology with cell cycle and epigenetic studies. Achieving a molecular understanding of key players such as cohesin is a requisite step for investigating how deterioration of these factors contributes to maternal age-dependent aneuploidy and infertility. The current trend towards advanced maternal age has increased the frequency of trisomic fetuses by 71% in the past ten years. Therefore, a better understanding of mammalian meiosis is relevant to human health.
How is sister chromatid cohesion maintained for months and decades in oocytes?
The inheritance of chromosomes from mother to daughter cell and from one generation to the next depends on sister chromatid cohesion mediated by the cohesin complex. Cohesin is especially important in meiosis, which is the specialized cell division giving rise to haploid gametes, egg and sperm. The paradigm of reproductive biology is that all oocytes are generated before birth. Cohesion is established during meiotic DNA replication, recombination occurs before birth, and oocytes remain arrested until ovulation triggers the first meiotic division several months (mouse) or decades (human) later. Does cohesin hold sister chromatids together for months and possibly decades without reinforcement? Alternatively, is cohesion reinforced during the long arrest? Using TEV protease technology which we pioneered in the mouse, molecular genetics, and 4D confocal live-cell imaging, we showed that no detectable cohesin turnover occurs in oocytes for several weeks (Figure 1). Our current work addresses the crucial question as to whether cohesion is reinforced during the months of arrest or, indeed, whether it is at all possible to generate cohesion after DNA replication (Figure 2). Future work will address the mechanisms that protect long-lived proteins such as cohesin, and what might go awry with age.
How is chromatin reprogramming coordinated with cell cycle progression in zygotes?
Fertilization triggers the second meiotic division and entry into the first embryonic cell cycle. During the zygote stage, maternal and paternal genomes remain as separate entities with distinct chromatin signatures. Maternal factors control sperm chromatin reorganization as protamines are replaced by histones and chromatin remodeling erases cell-type specific epigenetic marks. We are specifically interested in how chromatin organization, epigenetic reprogramming, and cell cycle progression are coordinated, which is currently poorly understood. This requires an interdisciplinary approach combining novel genome-wide methods to study chromatin organization with cell cycle kinetics documented by live-cell imaging.
We are testing candidate factors required for this process using conditional knockout mice. Knockout zygotes have the potential to be rescued by microinjection of mRNAs encoding target proteins. We are therefore developing this powerful system for in vivo structure-function studies in order to dissect the mechanisms of chromatin organization and cell cycle regulation in zygotes.