It is now approximately 25 years since the sheep Dolly, the first cloned mammal where the somatic cell nucleus from an adult donor was used for transfer, was born. So far, somatic cell nucleus transfer, where G1-phase nuclei are transferred into cytoplasts obtained by enucleation of mature metaphase II (MII) oocytes followed by the activation of the reconstructed cells, is the most efficient approach to reprogram/remodel the differentiated nucleus. In general, in an enucleated oocyte (cytoplast), the nuclear envelope (NE, membrane) of an injected somatic cell nucleus breaks down and chromosomes condense. This condensation phase is followed, after subsequent activation, by chromatin decondensation and formation of a pseudo-pronucleus (i) whose morphology should resemble the natural postfertilization pronuclei (PNs). Thus, the volume of the transferred nuclei increases considerably by incorporating the content released from the germinal vesicles (GVs). In parallel, the transferred nucleus genes must be reset and function similarly as the relevant genes in normal embryo reprogramming. This, among others, covers the relevant epigenetic modifications and the appropriate organization of chromatin in pseudo-pronuclei. While reprogramming in SCNT is often discussed, the remodeling of transferred nuclei is much less studied, particularly in the context of the developmental potential of SCNT embryos. It is now evident that correct reprogramming mirrors appropriate remodeling. At the same time, it is widely accepted that the process of rebuilding the nucleus following SCNT is instrumental to the overall success of this procedure. Thus, in our contribution, we will mostly focus on the remodeling of transferred nuclei. In particular, we discuss the oocyte organelles that are essential for the development of SCNT embryos.

• Keywords
• Nucleus, Remodeling, Reprogramming, Selective enucleation,
• MeSH
• Cell Nucleus metabolism   MeSH
• Chromatin metabolism   MeSH
• Oocytes MeSH
• Sheep genetics   MeSH
• Mammals genetics   MeSH
• Nuclear Transfer Techniques * veterinary   MeSH
• Animals MeSH
• Zygote * metabolism   MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Chromatin MeSH

Successful navigation of the mouse preimplantation stages of development, during which three distinct blastocyst lineages are derived, represents a prerequisite for continued development. We previously identified a role for p38-mitogen-activated kinases (p38-MAPK) regulating blastocyst inner cell mass (ICM) cell fate, specifically primitive endoderm (PrE) differentiation, that is intimately linked to rRNA precursor processing, polysome formation and protein translation regulation. Here, we develop this work by assaying the role of DEAD-box RNA helicase 21 (DDX21), a known regulator of rRNA processing, in the context of p38-MAPK regulation of preimplantation mouse embryo development. We show nuclear DDX21 protein is robustly expressed from the 16-cell stage, becoming exclusively nucleolar during blastocyst maturation, a localization dependent on active p38-MAPK. siRNA-mediated clonal Ddx21 knockdown within developing embryos is associated with profound cell-autonomous and non-autonomous proliferation defects and reduced blastocyst volume, by the equivalent peri-implantation blastocyst stage. Moreover, ICM residing Ddx21 knockdown clones express the EPI marker NANOG but rarely express the PrE differentiation marker GATA4. These data contribute further significance to the emerging importance of lineage-specific translation regulation, as identified for p38-MAPK, during mouse preimplantation development.

• Keywords
• DDX21, cell fate specification, p38-MAPK, preimplantation embryo development,
• MeSH
• Blastocyst cytology   metabolism   MeSH
• Cell Differentiation * genetics   MeSH
• Cell Lineage genetics   MeSH
• DEAD-box RNA Helicases genetics   metabolism   MeSH
• Embryonic Development * genetics   MeSH
• Fluorescent Antibody Technique MeSH
• Gene Knockdown Techniques MeSH
• p38 Mitogen-Activated Protein Kinases metabolism   MeSH
• Mice MeSH
• Signal Transduction MeSH
• Pregnancy MeSH
• Protein Transport MeSH
• Protein Binding MeSH
• Gene Expression Regulation, Developmental MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Pregnancy MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DDX21 protein, mouse MeSH Browser
• DEAD-box RNA Helicases MeSH
• p38 Mitogen-Activated Protein Kinases MeSH