Dynamic changes in epigenetic information underlie the transition of gametic chromatin to embryonic chromatin. These changes are thought to be required to initiate embryonic gene transcription and acquisition of pluripotency. Abnormalities in these transitions likely result in embryonic arrest at early preimplantation stages. Early embryonic mortality is a major cause of infertility in dairy cattle and humans. We and others established that histone methylation changes during preimplantation development is an active and enzymatically based process.
Specific Aim 1 will determine the locus-specific dynamics of histone methylation marks during preimplantation bovine development and their association with changes in gene expression. Using a rigorous experimental design, ChIP-seq and RNA-seq datasets across preimplantation bovine development will be generated to answer the following questions: 1) what is the role of repressive histone mark removal on EGA, 2) is deposition of activating histone marks related to EGA, 3) which genomic features escape histone modification remodeling, 4) what is the role of modified sperm histones for embryonic gene regulation, 5) how long does the asymmetric chromatin state persist and how does it affect embryonic gene expression, and 6) what is the epigenetic landscape of the totipotent state and how does it change with early lineage differentiation. Although it is generally accepted that the dramatic remodeling of gametic chromatin, driven by maternal factors, is required to initiate embryonic gene expression, the relationship between transcription and chromatin remodeling is poorly understood.
Specific Aim 2 will determine the relationship between embryonic gene expression and chromatin remodeling during bovine preimplantation development. The nature of transcripts produced by early embryos will be determined using full-length single-molecule RNA-sequencing and global chromatin accessibility landscapes interrogated using ATAC-seq. A rigorous experimental design using transcriptional inhibition and bioinformatics analysis will determine the characteristics of transcripts produced during embryo development and the relationship between transcription and chromatin remodeling. Together, these experiments will expand our understanding of basic mechanisms underlying embryonic development. This information will aid in developing diagnostic tools and interventions to treat infertility disorders. Moreover, understanding the transition from differentiated gametes to pluripotent blastomeres will shed light on mechanisms of chromatin remodeling of differentiated cells to pluripotency, which is relevant to cloning, stem cells, and cancer biology. Therefore, the aims of this project will contribute to improve our understanding of human development and health.
A greater understanding of biological mechanisms regulating epigenetic transformation during early embryonic development is critical to address problems of embryonic lethality observed in human infertility. Results from the proposed experiments will favorably impact the diagnosis and treatment of human fertility disorders, while simultaneously bettering foundational knowledge that can be applied to nuclear reprogramming methodologies necessary to generate pluripotent stem cells.