Understanding mechanism and improving efficiency of somatic cell nuclear transfer (SCNT) Abstract Understanding the mechanism of cell fate reprogramming is important for both basic biology and regenerative medicine. Of the currently available reprogramming techniques, somatic cell nuclear transfer (SCNT) is the only one that allows efficient and rapid reprogramming of terminally differentiated cells to the totipotent zygote-like state. Totipotency is the ability of a cell to give rise to an organism and its placental tissues. However, despite more than 50 years of effort since the first successful cloning by SCNT, very little progress has been made in understanding how SCNT reprogramming is achieved. Although ectopic expression of certain pluripotency transcription factors (TFs) can reprogram somatic cells into induced pluripotent stem cells (iPSCs), these cells are not totipotent. Moreover, accumulating evidence suggest that SCNT-mediated reprogramming is mechanistically different from that of transcription factor-based iPSC reprograming. Since maintaining undifferentiated stem cells in a lineage-unrestricted nave state is important for therapeutic purposes, understanding how differentiated somatic cells are reprogrammed into a totipotent state is of both biological and clinical importance. During SCNT-mediated reprogramming, donor cell genomes turn off their cell-type specific transcription programs and adopt a new gene expression profile that mimics that of totipotent zygotes. Our preliminary studies indicate that transcriptional reprogramming of donor cells is accomplished within 12 hours following SCNT, indicating that maternal factors present in oocytes can reset the chromatin state of somatic cells quickly upon nuclear transfer. Building upon this intriguing observation, as well as our recently developed techniques in analyzing chromatin accessibility of mouse zygotes and performing maternal factor depletion, we propose to understand the mechanism of SCNT reprogramming and improve SCNT efficiency with the following specific Aims: 1) Identifying and testing TFs and chromatin remodeling factors required for SCNT reprogramming; 2) Overcoming SCNT embryo developmental defects to increase animal term rate. Completion of the proposed study will not only identify oocyte factors important for SCNT-mediated reprogramming, but also improve the SCNT efficiency to achieve maximum term rate. These achievements will have far-reaching implications in the fields of development, stem cell, germ cell, chromatin biology, and regenerative medicine.

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Understanding mechanism and improving efficiency of somatic cell nuclear transfer (SCNT) Project Narrative Somatic cell nuclear transfer (SCNT) is the only reprogramming method to generate totipotent cells. However, very little is known about how SCNT is achieved. Building on two state-of-art techniques that we recently developed, we propose to dissect the mechanism of SCNT and improve its reprogramming efficiency.

National Institute of Health (NIH)
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
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Cellular, Molecular and Integrative Reproduction Study Section (CMIR)
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Ravindranath, Neelakanta
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Boston Children's Hospital
United States
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Djekidel, Mohamed Nadhir; Inoue, Azusa; Matoba, Shogo et al. (2018) Reprogramming of Chromatin Accessibility in Somatic Cell Nuclear Transfer Is DNA Replication Independent. Cell Rep 23:1939-1947
Matoba, Shogo; Wang, Huihan; Jiang, Lan et al. (2018) Loss of H3K27me3 Imprinting in Somatic Cell Nuclear Transfer Embryos Disrupts Post-Implantation Development. Cell Stem Cell 23:343-354.e5
Inoue, Azusa; Chen, Zhiyuan; Yin, Qiangzong et al. (2018) Maternal Eed knockout causes loss of H3K27me3 imprinting and random X inactivation in the extraembryonic cells. Genes Dev 32:1525-1536
Matoba, Shogo; Zhang, Yi (2018) Somatic Cell Nuclear Transfer Reprogramming: Mechanisms and Applications. Cell Stem Cell 23:471-485