Transcriptome Analyses Reveal Differential Transcriptional Profiles in Early- and Late-Dividing Porcine Somatic Cell Nuclear Transfer Embryos

Abstract

Somatic cell nuclear transfer (SCNT) is not only a valuable tool for understanding nuclear reprogramming, but it also facilitates the generation of genetically modified animals. However, the development of SCNT embryos has remained an uncontrollable process. It was reported that the SCNT embryos that complete the first cell division sooner are more likely to develop to the blastocyst stage, suggesting their better developmental competence. Therefore, to better understand the underlying molecular mechanisms, RNA-seq of pig SCNT embryos that were early-dividing (24 h postactivation) and late-dividing (36 h postactivation) was performed. Our analysis revealed that early- and late-dividing embryos have distinct RNA profiles, and, in all, 3077 genes were differentially expressed. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that early-dividing embryos exhibited higher expression in genes that participated in the meiotic cell cycle, while enrichment of RNA processing- and translation-related genes was found in late-dividing embryos. There are also fewer somatic memory genes such as FLRT2, ADAMTS1, and FOXR1, which are abnormally activated or suppressed in early-dividing cloned embryos. These results show that early-dividing SCNT embryos have different transcriptional profiles than late-dividing embryos. Early division of SCNT embryos may be associated with their better reprogramming capacity, and somatic memory genes may act as a reprogramming barrier in pig SCNT reprogramming.

RNA-Seq Reveals the Underlying Molecular Mechanism of First Cleavage Time Affecting Porcine Embryo Development

“…It is uncertain whether DNA damage causes protein misfolding in the ER. It has been reported that 1896 DEGs were identified between SCNT early cleavage embryos and late cleavage embryos [ 19 ]. Similar results to ours showed that the DEGs were enriched in the ribosome, mRNA surveillance, protein processing in the endoplasmic reticulum, and spliceosome pathways [ 19 ].…”

“…Diverse studies have found that relatively early cleavage embryos relation to better developmental competence than those cleaving later embryos in pig [ 11 , 12 ], human [ 13 ], buffalo [ 14 ], cattle [ 7 , 15 ], goat [ 16 ], hamster [ 17 ], and mouse [ 18 ]. It has been reported that early cleavage embryos exhibited higher expression in genes that participated in the meiotic cell cycle, while enrichment of RNA processing- and translation-related genes was found in porcine late cleavage embryos of somatic cell nuclear transfer (SCNT) [ 19 ]. While the delay of cell division might be a consequence of chromosomal aberrations and DNA damage [ 20 ], slow cleaving embryos have a higher caspase activity in comparison to fast cleavers [ 21 ].…”

Epigenetic manipulation to improve mouse SCNT embryonic development

“…In 2016, Liu et al (2020) found that overexpression of Kdm5b , an H3K4me3 demethylase, rescued 4-cell arrest in mouse cloned embryos. Moreover, the combination of K4m4b and Kdm5b improved the SCNT blastocyst rate to over 95%, and 11.1% of reconstructed embryos developed to full term.…”

“…The efficiency of SCNT-based cloning in mammals remains at relatively or extremely low levels, and it appears to differ in a species-specific manner. To improve the SCNT efficiency, extensive efforts are needed to precisely identify the epigenetic factors and molecular mechanisms involved in the reprogramming of the nuclear donor cell’s (NDC’s) genome in somatic cell cloned embryos ( Cao et al, 2020 ; Liu et al, 2020 ; Feng et al, 2022 ; Li et al, 2022 ). Thus far, the provenance and type of NDCs have been shown to influence the capability of donor genomic DNA to be epigenetically reprogrammed in the oocyte cytoplasm during SCNT ( Samiec and Skrzyszowska, 2010 , 2013 ; Sadeesh et al, 2016 ; Wiater et al, 2021b ; Xu et al, 2021 ; Zhang et al, 2022 ).…”

Enhancement of Chromatin and Epigenetic Reprogramming in Porcine SCNT Embryos—Progresses and Perspectives

“…Proper reprogramming to totipotency in SCNT embryos can be hampered by both lack of gene activation and failure to inactivate the transcriptional memory of somatic nuclei following nuclear transfer ( Ng and Gurdon, 2005 ; Matoba et al, 2014 ). The transcriptional memory of somatic cells was more efficiently reprogrammed in SCNT embryos that cleaved early and produced higher blastocyst rates than late cleaving embryos ( Liu et al, 2020 ). In addition, attenuation of the transcriptional memory for 15 h after nuclear transfer, using the inhibitor of RNA polymerase II, 5,6-Dichlorobenzimidazole 1-β- d -ribofuranoside (DRB), improved gene expression in bovine SCNT embryos and increased blastocyst cell numbers ( Rissi et al, 2018 ).…”