Abstract
Somatic cell nuclear transfer (SCNT) is a very powerful technique used to produce genetically identical or modified animals. However, the cloning efficiency in mammals remains low. In this study, we aimed to explore the effects of vitamin C (Vc)-treated donor cells on cloned embryos. As a result, Vc treatment relaxed the chromatin of donor cells and improved cloned embryo development. RNA sequencing was adopted to investigate the changes in the transcriptional profiles in early embryos. We found that Vc treatment increased the expression of genes involved in the cell–substrate adherens junction. Gene ontology (GO) analysis revealed that Vc treatment facilitated the activation of autophagy, which was deficient in cloned two-cell embryos. Rapamycin, an effective autophagy activator, increased the formation of cloned blastocysts (36.0% vs. 25.6%, p < 0.05). Abnormal expression of some coding genes and long non-coding RNAs in cloned embryos was restored by Vc treatment, including the zinc-finger protein 641 (ZNF641). ZNF641 compensation by means of mRNA microinjection improved the developmental potential of cloned embryos. Moreover, Vc treatment rescued some deficient RNA-editing sites in cloned two-cell embryos. Collectively, Vc-treated donor cells improved the development of the cloned embryo by affecting embryonic transcription. This study provided useful resources for future work to promote the reprogramming process in SCNT embryos.
Highlights
Animal cloning has been successful since 1997 when “Dolly the sheep” was first created by nuclear transfer using differentiated somatic cells (SCNT) [1]
The cleavage of cloned embryos was slower than In vitro fertilization (IVF) embryos on day 1 (51.8% vs 68.5%, p < 0.05) and was accelerated using Vitamin C (Vc)-treated donor cells (65.0% vs 51.8%)
We had found that Vc treatment decreased DNA methylation in donor cells; we speculated that Vc treatment might relax chromatin of donor cells
Summary
Animal cloning has been successful since 1997 when “Dolly the sheep” was first created by nuclear transfer using differentiated somatic cells (SCNT) [1]. Determining the underlying mechanism of genetic reprogramming to improve the nuclear transfer (NT) procedures and optimize the outcomes of SCNT has proven to be difficult. In fertilized embryos, a series of reprogramming events occur efficiently. Incomplete nuclear reprogramming is the most serious problem in cloned embryos [5,6]. During the production of SCNT embryos, the epigenetic memory of donor somatic cells must be erased to ensure appropriated embryonic access to the genome. The donor cells are highly differentiated and difficult to radically reprogram into a pluripotent state through epigenetic modifications [7]. Drugs that modify and remold the epigenome have been widely studied in the context of SCNT
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