Heterogeneity in Somatic Cell Nuclear Transfer Embryo Reprogramming Across Mammalian Species
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ABSTRACT: Somatic cell nuclear transfer (SCNT) enables the reprogramming of terminally differentiated somatic cells into a totipotent state, yet whether this process is governed by conserved mechanisms across mammalian species remains poorly understood. Here, we employed low input transcriptomics to profile global transcriptional dynamics during zygotic genome activation in SCNT embryos from five species: mouse, pig, cattle, goat, and sheep. Our analysis revealed distinct genome-wide expression patterns among these species, with 70.17% of differentially expressed genes (DEGs) being species-specific, while only 9.17% were shared DEGs. Functional annotation of the shared DEGs highlighted their enrichment in processes such as mRNA transcription, translation, and carbohydrate metabolism. Notably, we observed widespread pathway overactivation in SCNT embryos from cattle, mouse, and pig, whereas goat and sheep SCNT embryos exhibited broad suppression. Furthermore, weighted gene co-expression network analysis (WGCNA) demonstrated that species-specific effects exerted a far greater influence on reprogramming outcomes than the method of embryo generation (Fertilization or SCNT). Through systematic identification of key transcription factors, signaling pathways, epigenetic markers, and alternative splicing events, we uncovered species-specific regulatory patterns underlying reprogramming. Additionally, genome browser analysis revealed regional chromosomal expression anomalies in GDF9 and BMP15 during reprogramming across species. Collectively, our findings provide critical insights into the divergent mechanisms of reprogramming in mammalian species and establish a robust theoretical foundation for future studies in this field.
Project description:We carried out a cross species cattle-sheep array comparative genome hybridization (aCGH) experiment in order to identify copy number variations (CNVs) in the sheep genome analysing animals of Italian dairy breeds (Sarda, Bagnolese, Laticauda, Massese and Valle del Belice) using a tiling oligonucleotide array with ~385,000 probes designed on the bovine genome. We identified 135 CNV regions (CNVRs) covering about 10.5 Mb of the virtual sheep genome referred to the bovine genome (0.398%) with a mean and median equal to 77.6 kb and 55.9 kb, respectively. A comparative analysis between the identified sheep CNVRs and those reported in the cattle and goat genomes indicated that overlaps between sheep and goat and sheep and cattle CNVRs are highly significant (P<0.0001) suggesting that several chromosome regions might contain recurrent interspecies CNVRs. Many sheep CNVs affect genes with important biological functions. Further studies are needed to evaluate the functional relevance of these CNVs.
Project description:We carried out a cross species cattle-sheep array comparative genome hybridization (aCGH) experiment in order to identify copy number variations (CNVs) in the sheep genome analysing animals of Italian dairy breeds (Sarda, Bagnolese, Laticauda, Massese and Valle del Belice) using a tiling oligonucleotide array with ~385,000 probes designed on the bovine genome. We identified 135 CNV regions (CNVRs) covering about 10.5 Mb of the virtual sheep genome referred to the bovine genome (0.398%) with a mean and median equal to 77.6 kb and 55.9 kb, respectively. A comparative analysis between the identified sheep CNVRs and those reported in the cattle and goat genomes indicated that overlaps between sheep and goat and sheep and cattle CNVRs are highly significant (P<0.0001) suggesting that several chromosome regions might contain recurrent interspecies CNVRs. Many sheep CNVs affect genes with important biological functions. Further studies are needed to evaluate the functional relevance of these CNVs. In this study we made use of the high conservation and homology between the cattle and sheep genomes determined by their phylogenetic closeness to identify CNVs in sheep applying the same approach we carried out to identify CNVs in the goat genome. We used a custom tiling array including ~385,000 oligonucleotide probes designed on the Btau_4.0 version of the Bos taurus genome assembly and analysed genomic DNA samples of 11 sheep belonging to 6 different Italian dairy sheep breeds (2 Sarda, 2 Bagnolese, 2 Comisana, 2 Massese, 2 Laticauda and 1 Valle del Belice) compared to the reference DNA of another Sarda sheep.
Project description:Pig cloning by somatic cell nuclear transfer (SCNT) frequently undergoes incomplete epigenetic remodeling during the maternal-to-zygotic transition, which leads to a significant embryonic loss before implantation. Here, we generated the first genome-wide landscapes of histone methylation in pig SCNT embryos. Excessive H3K9me3 and H3K27me3, but not H3K4me3, were observed in the genomic regions with unfaithful embryonic genome activation and donor cell-specific gene silencing. A combination of H3K9 demethylase KDM4A and GSK126, an inhibitor of H3K27me3 writer, could remove these epigenetic barriers and restore the global transcriptome in SCNT embryos. More importantly, TDG was defined as a pig-specific epigenetic regulator for nuclear reprogramming, which was not reactivated by H3K9me3 and H3K27me3 removal. Both combined treatment and transient TDG overexpression could promote DNA demethylation and enhance the blastocyst forming rates of SCNT embryos, which offers valuable methods to increase the cloning efficiency of genome-edited pigs for agricultural and biomedical purposes.
Project description:Pig cloning by somatic cell nuclear transfer (SCNT) frequently undergoes incomplete epigenetic remodeling during the maternal-to-zygotic transition, which leads to a significant embryonic loss before implantation. Here, we generated the first genome-wide landscapes of histone methylation in pig SCNT embryos. Excessive H3K9me3 and H3K27me3, but not H3K4me3, were observed in the genomic regions with unfaithful embryonic genome activation and donor cell-specific gene silencing. A combination of H3K9 demethylase KDM4A and GSK126, an inhibitor of H3K27me3 writer, could remove these epigenetic barriers and restore the global transcriptome in SCNT embryos. More importantly, TDG was defined as a pig-specific epigenetic regulator for nuclear reprogramming, which was not reactivated by H3K9me3 and H3K27me3 removal. Both combined treatment and transient TDG overexpression could promote DNA demethylation and enhance the blastocyst forming rates of SCNT embryos, which offers valuable methods to increase the cloning efficiency of genome-edited pigs for agricultural and biomedical purposes.
Project description:Pig cloning by somatic cell nuclear transfer (SCNT) frequently undergoes incomplete epigenetic remodeling during the maternal-to-zygotic transition, which leads to a significant embryonic loss before implantation. Here, we generated the first genome-wide landscapes of histone methylation in pig SCNT embryos. Excessive H3K9me3 and H3K27me3, but not H3K4me3, were observed in the genomic regions with unfaithful embryonic genome activation and donor cell-specific gene silencing. A combination of H3K9 demethylase KDM4A and GSK126, an inhibitor of H3K27me3 writer, could remove these epigenetic barriers and restore the global transcriptome in SCNT embryos. More importantly, TDG was defined as a pig-specific epigenetic regulator for nuclear reprogramming, which was not reactivated by H3K9me3 and H3K27me3 removal. Both combined treatment and transient TDG overexpression could promote DNA demethylation and enhance the blastocyst forming rates of SCNT embryos, which offers valuable methods to increase the cloning efficiency of genome-edited pigs for agricultural and biomedical purposes. This SuperSeries is composed of the SubSeries listed below.
Project description:Pig cloning by somatic cell nuclear transfer (SCNT) frequently undergoes incomplete epigenetic remodeling during the maternal-to-zygotic transition, which leads to a significant embryonic loss before implantation. Here, we generated the first genome-wide landscapes of histone methylation in pig SCNT embryos. Excessive H3K9me3 and H3K27me3, but not H3K4me3, were observed in the genomic regions with unfaithful embryonic genome activation and donor cell-specific gene silencing. A combination of H3K9 demethylase KDM4A and GSK126, an inhibitor of H3K27me3 writer, could remove these epigenetic barriers and restore the global transcriptome in SCNT embryos. More importantly, TDG was defined as a pig-specific epigenetic regulator for nuclear reprogramming, which was not reactivated by H3K9me3 and H3K27me3 removal. Both combined treatment and transient TDG overexpression could promote DNA demethylation and enhance the blastocyst forming rates of SCNT embryos, which offers valuable methods to increase the cloning efficiency of genome-edited pigs for agricultural and biomedical purposes.
Project description:Purpose: High-altitude adaptive evolution of transcription, and the convergence and divergence of transcriptional alteration across species in response to high-altitude environments, is an important topic of broad interest to the general biology community. Our study aims to answer this important biological question. Methods: We generated deep transcriptome data of high- and low- altitude populations across four species: chicken, pig, goat and sheep, as well as high-altitude yak and low-altitude cattle, from six tissues (heart, kidney, liver, lung, skeletal muscle and spleen). Results: Here we provide a comprehensive comparative transcriptome landscape of expression and alternative splicing variation between low- and high-altitude populations across multiple species for distinct tissues. Conclusions: Our data serves a valuable resource for further study on adaptive transcription evolution and identification of candidate adaptive genes.
Project description:Somatic cell nuclear transplantation (SCNT) can transform highly differentiated donor nuclei into pluripotent nuclei through the large-scale reprogramming of chromatin. The reprogramming of chromatin has been documented to take place in the first few hours after SCNT embryo activation. Thus, studies that characterize dynamic changes in chromatin during the first few hours after embryo activation could provide insight into the mechanism and significance of genome-wide reprogramming. However, few studies have examined the epigenetic remodeling of reconstructed embryos during the early stage of reprogramming. Here, we conducted ATAC-seq on 50 porcine SCNT-HMC embryos and 50 parthenogenetic activation (PA) embryos 10 h after activation. Along with pig embryonic fibroblast (PEF) ATAC-seq data, we found low levels of chromatin accessibility and gene transcription in SCNT and PA embryos. Moreover, PEF genes and the X chromosome became inaccessible during embryo reprogramming. GO enrichment analysis revealed that the molecular functions related to accessible chromatin in embryos primarily included transcriptional regulatory activity and SMAD binding. The differentially accessible chromatin sites between SCNT and PEF were primarily related to transcriptional activity and histone modification. These results indicated that despite the tight chromatin structure during the early stage of embryo reprogramming, some accessible chromatin sites, which were primarily distributed in the intergenic region, were still detected. Dynamic changes in chromatin accessibility during reprogramming were primarily related to transcriptional activity and histone modification. Generally, this study provided new insight into the dynamics and importance of chromatin accessibility during the early stages of embryo reprogramming.
Project description:MicroRNAs (miRNAs) are small non-coding RNAs that regulate the post transcriptional control of several pathway intermediates, and essential for regulation in skeletal muscle of many species, such as mice, cattle, pig and so on. However, a little number of miRNAs have been reported in the muscle development of goat. In this study, the longissimus dorsi transcripts of goat at 1- and 10-month-old were analyzed for RNA-seq and miRNA-seq. The results showed that 10-month-old Longlin goat expressed 327 up- and 419 down-regulated differentially expressed genes (DEGs) compared with the 1-month-old were founded. In addition, 20 co-up-regulated and 55 co-down-regulated miRNAs involved in muscle fiber hypertrophy of goat were identified in 10-month-old Longlin and Nubian goat compared with 1-month-old. Five miRNA–mRNA pairs (chi-let-7b-3p-MIRLET7A, chi-miR193b-3p-MMP14, chi-miR-355-5p-DGAT2, novel_128-LOC102178119, novel_140-SOD3) involved in the goat skeletal muscle development were identified by miRNA–mRNA negative correlation network analysis. Our results provided an insight into the functional roles of miRNAs of goat muscle-associated miRNAs, allowing us to better understand the transformation of miRNA roles during mammalian muscle development.