Epitranscriptomic regulation and genome stability in meiosis and fertilization

Epitranscriptomic regulation and genome stability in meiosis and fertilizationOur kajor focus is to establish low-input and single cell strategies for analysing the dynamics of RNA modifications in meiosis and early embryo development. Single-cell analysis will also enable important studies on cancer heterogeneity, for which we focus on bladder cancer.In the last year we have succeded in providing high quality data on the dynamics on 6-methyladenine (m6A) in single oocytes and zygotes, as well as in limited (20) number of somatic cells. Two studies has been published: Li, Wang et al., Single-cell m6A mapping in vivo using picoMeRIP-seq. Nat Biotechnol. 2023 Wang, Li et al., The RNA m6A landscape of mouse oocytes and preimplantation embryos. Nat Struct Mol Biol. 2023 We are currently completing a study on human oocytes and preimplantation embryos for comparison with murine samples. Additionally, we study embryos arrested at the 2 and 2-cell stage. In future studies (application for funding elsewhere), this method will be used for analysing cancer heterogeneity

Epitranscriptomic regulation and genome stability in meiosis and fertilizationMeiosis is believed to have developed very early in eukaryotic evolution and is a key component of sexual reproduction and the equal delivery and recombining of maternal and paternal chromosomes. We here study in detail the role of dynamic and reversible methylations in mRNA during meiosis and in the preimplantation embryo.Meiosis is believed to have developed very early in eukaryotic evolution and is a key component of sexual reproduction and the equal delivery and recombining of maternal and paternal chromosomes. One of the most profound cellular changes in the life of an organism is the remarkable reprogramming of mRNA transcripts in the early preimplantation embryo. The reliance on presynthesized mRNAs is a unique feature of meiosis (Susor et al., 2016) and shortly after fertilization, embryos cross a stage during which developmental control is handed from maternally supplied mRNAs from the mature oocyte to mRNAs transcribed from the zygote; the maternal-to-zygotic transition (MZT). We here propose to study in detail the role of dynamic and reversible methylations in mRNA in meiosis and through the MZT in the preimplantation embryo. The project will focus on single-cell and single embryo analysis. Our most recent discovery of epitranscriptomic regulation of R-loop structures will be pursued to elucidate its role in genome (in)stability and for the potential regulation of mutational load of meiotic cells. During 2022 we have initiated the characterization of the epitranscriptome in stage-specific single cells during female meiosis. We have also preliminary data on the epitranscriptome of human oocytes and preimplantation embryos as well as in mutant oocytes from mice lacking a demethylase that removes one of the key modification referred to as epitranscriptomic modification. These two studies are expected to be submitted for publication summer/autumn 2023. One manuscript is accepted for publication (corresponding authors Arne Klungland, John Arne Dahl and Kin Fai Au (last): Yunhao Wang Yanjiao L et al, The RNA m6A landscape in mouse oocytes and preimplantation embryos, Nat Struct Mol Biol in press and another being resubmitted following the request of minor 2. Revision (corresponding authors Arne Klungland, Kin Fai Au and John Arne Dahl (last): Yanjiao Li, Yunhao Wang et al, Single-cell MeRIP-seq maps m6A in mouse oocytes and embryos (2. Revision Nat Biotech)

Epitranscriptomic regulation and genome stability in meiosis and fertilizationMeiosis is believed to have developed very early in eukaryotic evolution and is a key component of sexual reproduction and the equal delivery and recombining of maternal and paternal chromosomes. We here study in detail the role of dynamic and reversible methylations in mRNA during meiosis and in the preimplantation embryo.Meiosis is believed to have developed very early in eukaryotic evolution and is a key component of sexual reproduction and the equal delivery and recombining of maternal and paternal chromosomes. One of the most profound cellular changes in the life of an organism is the remarkable reprogramming of mRNA transcripts in the early preimplantation embryo. The reliance on presynthesized mRNAs is a unique feature of meiosis (Susor et al., 2016) and shortly after fertilization, embryos cross a stage during which developmental control is handed from maternally supplied mRNAs from the mature oocyte to mRNAs transcribed from the zygote; the maternal-to-zygotic transition (MZT). We here propose to study in detail the role of dynamic and reversible methylations in mRNA in meiosis and through the MZT in the preimplantation embryo. The project will focus on single-cell and single embryo analysis. Our most recent discovery of epitranscriptomic regulation of R-loop structures will be pursued to elucidate its role in genome (in)stability and for the potential regulation of mutational load of meiotic cells. Methylations of DNA and histone residues regulate transcription and the discoveries of demethylases that remove these marks led to a tremendous progress in our understanding of gene regulation. While post-transcriptional RNA modifications were identified several decades ago, their reversible nature has only recently been described. Our studies will focus on defining the temporal and spatial activities of m6A readers and erasers in meiosis. Owing to technological advances, knowledge of epitranscriptomic marks and their writers, readers and erasers has recently advanced tremendously. The project will combine our expertise and contribution to the discovery of reversible epitranscriptomic marks and studies on the epigenetic regulation of the MZT with our previous and studies on DNA stability. Aylin Cengiz, PhD candidate funded by the project. Her specific role in the project is: 1. Study the consequence of disrupted m6A regulation for progression of oocytes through meiosis I 2. Genome stability in oocytes and early embryos chemically stimulated for progression through meiosis I and cellular division. 3. Chromosomal damage to DNA in the nucleus and polar body. Kangxuan Jin, Researcher funded by the project. His specific role in the project is: 1. Characterize in detail the m6A transcriptome in meiotic oocytes lacking ALKBH5. ALKBH5 is a m6A demethylase 2. m6A dependent expression of retrotransposons in meiosis and the early embryo. Aylin Cengiz and Kangxuan are working closely together. Additionally, we have already obtained results from Melina Schuh (collaborator Gøttingen) relating to microtubuli formation and chromosome misalignment and during mouse oocyte meiosison (SN and NSN oocytes in meiosis I) relating to m6A deficiency due to ALKBH5 deletion.