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Epigenetic integrity of paternal imprints enhances the developmental potential of androgenetic haploid embryonic stem cells.

ABSTRACT: The use of two inhibitors of Mek1/2 and Gsk3β (2i) promotes the generation of mouse diploid and haploid embryonic stem cells (ESCs) from the inner cell mass of biparental and uniparental blastocysts, respectively. However, a system enabling long-term maintenance of imprints in ESCs has proven challenging. Here, we report that the use of a two-step a2i (alternative two inhibitors of Src and Gsk3β, TSa2i) derivation/culture protocol results in the establishment of androgenetic haploid ESCs (AG-haESCs) with stable DNA methylation at paternal DMRs (differentially DNA methylated regions) up to passage 60 that can efficiently support generating mice upon oocyte injection. We also show coexistence of H3K9me3 marks and ZFP57 bindings with intact DMR methylations. Furthermore, we demonstrate that TSa2i-treated AG-haESCs are a heterogeneous cell population regarding paternal DMR methylation. Strikingly, AG-haESCs with late passages display increased paternal-DMR methylations and improved developmental potential compared to early-passage cells, in part through the enhanced proliferation of H19-DMR hypermethylated cells. Together, we establish AG-haESCs that can long-term maintain paternal imprints.

SUBMITTER: Zhang H

PROVIDER: S-EPMC8783938 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Epigenetic integrity of paternal imprints enhances the developmental potential of androgenetic haploid embryonic stem cells

Project description:The use of two inhibitors of Mek1/2 and Gsk3β (2i) promotes the generation of mouse diploid and haploid embryonic stem cells (ESCs) from the inner cell mass of biparental and uniparental blastocysts, respectively. However, a system enabling long-term maintenance of imprints in ESCs has proven challenging. Here, we report that usage of a two-step a2i (alternative two inhibitors of Src and Gsk3β, TSa2i) derivation/culture protocol results in the establishment of androgenetic haploid ESCs (AG-haESCs) with stable DNA methylation at paternal DMRs (differentially DNA methylated regions) up to passage 60 that can efficiently support generating mice upon oocyte injection. We also show coexistence of H3K9me3 marks and ZFP57 bindings with intact DMR methylations. Furthermore, we demonstrate that TSa2i-treated AG-haESCs are a heterogeneous cell population regarding paternal DMR methylation. Strikingly, AG-haESCs with late passages display increased paternal-DMR methylations and improved developmental potential compared to early-passage cells, in part through the enhanced proliferation of H19-DMR hypermethylated cells. Together, we establish AG-haESCs that can long-term maintain paternal imprints.

2022-01-26 | GSE168447 | GEO

Epigenetic integrity of paternal imprints enhances the developmental potential of androgenetic haploid embryonic stem cells

Project description:Epigenetic integrity of paternal imprints enhances the developmental potential of androgenetic haploid embryonic stem cells

| PRJNA707435 | ENA

Derivation of Haploid Neurons from Mouse Androgenetic Haploid Embryonic Stem Cells.

Project description: Not available

| S-EPMC5567516 | biostudies-literature

Transcriptome profiles and chromatin states in mouse androgenetic haploid embryonic stem cells.

Project description:Haploid embryonic stem cells (haESCs) are derived from the inner cell mass of the haploid blastocyst, containing only one set of chromosomes. Extensive and accurate chromatin remodelling occurs during haESC derivation, but the intrinsic transcriptome profiles and chromatin structure of haESCs have not been fully explored. We profiled the transcriptomes, nucleosome positioning, and key histone modifications of four mouse haESC lines, and compared these profiles with those of other closely-related stem cell lines, MII oocytes, round spermatids, sperm, and mouse embryonic fibroblasts. haESCs had transcriptome profiles closer to those of naïve pluripotent stem cells. Consistent with the one X chromosome in haESCs, Xist was repressed, indicating no X chromosome inactivation. haESCs and ESCs shared a similar global chromatin structure. However, a nucleosome depletion region was identified in 2056 promoters in ESCs, which was absent in haESCs. Furthermore, three characteristic spatial relationships were formed between transcription factor motifs and nucleosomes in both haESCs and ESCs, specifically in the linker region, on the nucleosome central surface, and nucleosome borders. Furthermore, the chromatin state of 4259 enhancers was off in haESCs but active in ESCs. Functional annotation of these enhancers revealed enrichment in regulation of the cell cycle, a predominantly reported mechanism of haESC self-diploidization. Notably, the transcriptome profiles and chromatin structure of haESCs were highly preserved during passaging but different from those of differentiated cell types.

| S-EPMC10472531 | biostudies-literature

Androgenetic haploid embryonic stem cells

Project description:This SuperSeries is composed of the SubSeries listed below.

2012-08-30 | GSE39392 | GEO

Rapidly generating knockout mice from H19-Igf2 engineered androgenetic haploid embryonic stem cells.

Project description:Haploid mammalian embryonic stem cells (ESCs) hold great promise for functional genetic studies and assisted reproduction. Recently, rodent androgenetic haploid ESCs (AG-haESCs) were generated from androgenetic blastocysts and functioned like sperm to produce viable offspring via the intracytoplasmic AG-haESCs injection into oocytes. However, the efficiency of this reproduction was very low. Most pups were growth-retarded and died shortly after birth, which is not practical for producing knockout animals. Further investigation suggested a possible link between the low birthrate and aberrant expression of imprinted genes. Here, we report the high-frequency generation of healthy, fertile mice from H19-Igf2 imprinting-locus modified AG-haESCs, which maintained normal paternal imprinting and pluripotency. Moreover, it is feasible to perform further genetic manipulations in these AG-haESCs. Our study provides a reliable and efficient tool to rapidly produce gene-modified mouse models and will benefit reproductive medicine in the future.

| S-EPMC4860787 | biostudies-literature

CGH analysis of androgenetic haploid embryonic stem cells

2012-08-30 | GSE39390 | GEO

CGH analysis of androgenetic haploid embryonic stem cells

Project description:Comparative genomic hybridization (CGH) analysis of androgenetic haploid embryonic stem cells was done with 129/Sv male mouse kidney genomic DNA as reference. The results revealed no consistent copy number variations (CNVs) among the examined cell lines (AH129-5, AH129-N1 and AH129-NC1), except for the global difference on the X chromosome to the male control. Genomic DNA of FACS sorted G0/G1 stage ahES cells was compared with 129/Sv male mouse kidney genomic DNA reference by the NimbleGen Mouse CGH 3x720K Whole-Genome Tiling Array (Build MM9) [100718_MM9_WG_CGH_HX3]

2012-08-30 | E-GEOD-39390 | biostudies-arrayexpress

CGH analysis of AG-haESCs (androgenetic haploid embryonic stem cells)

Project description:The genomic DNA sample of AG-haESCs were compared to the C57BL/6J male mouse kidney by comparative genomic hybridization. The data confirmed that the haploid cells sustained genome integrity.

2012-04-27 | GSE35786 | GEO

Reduced Self-Diploidization and Improved Survival of Semi-cloned Mice Produced from Androgenetic Haploid Embryonic Stem Cells through Overexpression of Dnmt3b.

Project description:Androgenetic haploid embryonic stem cells (AG-haESCs) hold great promise for exploring gene functions and generating gene-edited semi-cloned (SC) mice. However, the high incidence of self-diploidization and low efficiency of SC mouse production are major obstacles preventing widespread use of these cells. Moreover, although SC mice generation could be greatly improved by knocking out the differentially methylated regions of two imprinted genes, 50% of the SC mice did not survive into adulthood. Here, we found that the genome-wide DNA methylation level in AG-haESCs is extremely low. Subsequently, downregulation of both de novo methyltransferase Dnmt3b and other methylation-related genes was determined to be responsible for DNA hypomethylation. We further demonstrated that ectopic expression of Dnmt3b in AG-haESCs could effectively improve DNA methylation level, and the high incidence of self-diploidization could be markedly rescued. More importantly, the developmental potential of SC embryos was improved, and most SC mice could survive into adulthood.

| S-EPMC5831042 | biostudies-literature

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