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Aberrant nucleosome organization in mouse SCNT embryos revealed by ULI-MNase-seq.

ABSTRACT: Somatic cell nuclear transfer (SCNT) can reprogram terminally differentiated somatic cells into totipotent embryos, but with multiple defects. The nucleosome positioning, as an important epigenetic regulator for gene expression, is largely unexplored during SCNT embryonic development. Here, we mapped genome-wide nucleosome profiles in mouse SCNT embryos using ultra-low-input MNase-seq (ULI-MNase-seq). We found that the nucleosome-depleted regions (NDRs) around promoters underwent dramatic reestablishment, which is consistent with the cell cycle. Dynamics of nucleosome position in SCNT embryos were delayed compared to fertilized embryos. Subsequently, we found that the aberrant gene expression levels in inner cell mass (ICM) were positively correlated with promoter NDRs in donor cells, which indicated that the memory of nucleosome occupancy in donor cells was a potential barrier for SCNT-mediated reprogramming. We further confirmed that the histone acetylation level of donor cells was associated with the memory of promoter NDRs. Our study provides insight into nucleosome reconfiguration during SCNT preimplantation embryonic development.

SUBMITTER: Yang L

PROVIDER: S-EPMC9287678 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature

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Aberrant nucleosome organization in mouse SCNT embryos revealed by ULI-MNase-seq.

Yang Lingyue L Xu Xiaocui X Xu Ruimin R Chen Chuan C Zhang Xiaolei X Chen Mo M Kou Xiaochen X Zhao Yanhong Y Wang Hong H Liu Xiaoyu X Gao Shaorong S Li Chong C

Stem cell reports 20220623 7

Somatic cell nuclear transfer (SCNT) can reprogram terminally differentiated somatic cells into totipotent embryos, but with multiple defects. The nucleosome positioning, as an important epigenetic regulator for gene expression, is largely unexplored during SCNT embryonic development. Here, we mapped genome-wide nucleosome profiles in mouse SCNT embryos using ultra-low-input MNase-seq (ULI-MNase-seq). We found that the nucleosome-depleted regions (NDRs) around promoters underwent dramatic reesta ...[more]

PMID: 35750045

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Project description:BackgroundTFIIS stimulates RNA cleavage by RNA polymerase II and promotes the resolution of backtracking events. TFIIS acts in the chromatin context, but its contribution to the chromatin landscape has not yet been investigated. Co-transcriptional chromatin alterations include subtle changes in nucleosome positioning, like those expected to be elicited by TFIIS, which are elusive to detect. The most popular method to map nucleosomes involves intensive chromatin digestion by micrococcal nuclease (MNase). Maps based on these exhaustively digested samples miss any MNase-sensitive nucleosomes caused by transcription. In contrast, partial digestion approaches preserve such nucleosomes, but introduce noise due to MNase sequence preferences. A systematic way of correcting this bias for massively parallel sequencing experiments is still missing.ResultsTo investigate the contribution of TFIIS to the chromatin landscape, we developed a refined nucleosome-mapping method in Saccharomyces cerevisiae. Based on partial MNase digestion and a sequence-bias correction derived from naked DNA cleavage, the refined method efficiently mapped nucleosomes in promoter regions rich in MNase-sensitive structures. The naked DNA correction was also important for mapping gene body nucleosomes, particularly in those genes whose core promoters contain a canonical TATA element. With this improved method, we analyzed the global nucleosomal changes caused by lack of TFIIS. We detected a general increase in nucleosomal fuzziness and more restricted changes in nucleosome occupancy, which concentrated in some gene categories. The TATA-containing genes were preferentially associated with decreased occupancy in gene bodies, whereas the TATA-like genes did so with increased fuzziness. The detected chromatin alterations correlated with functional defects in nascent transcription, as revealed by genomic run-on experiments.ConclusionsThe combination of partial MNase digestion and naked DNA correction of the sequence bias is a precise nucleosomal mapping method that does not exclude MNase-sensitive nucleosomes. This method is useful for detecting subtle alterations in nucleosome positioning produced by lack of TFIIS. Their analysis revealed that TFIIS generally contributed to nucleosome positioning in both gene promoters and bodies. The independent effect of lack of TFIIS on nucleosome occupancy and fuzziness supports the existence of alternative chromatin dynamics during transcription elongation.

Project description:Nucleosome positioning is critical to chromatin accessibility and is associated with gene expression programs in cells1-3. Previous nucleosome mapping methods assemble profiles from cell populations and reveal a cell-averaged pattern: nucleosomes are positioned and form a phased array that surrounds the transcription start sites of active genes3-6 and DNase I hypersensitive sites7. However, even in a homogenous population of cells, cells exhibit heterogeneity in expression in response to active signalling8,9 that may be related to heterogeneity in chromatin accessibility10-12. Here we report a technique, termed single-cell micrococcal nuclease sequencing (scMNase-seq), that can be used to simultaneously measure genome-wide nucleosome positioning and chromatin accessibility in single cells. Application of scMNase-seq to NIH3T3 cells, mouse primary naive CD4 T cells and mouse embryonic stem cells reveals two principles of nucleosome organization: first, nucleosomes in heterochromatin regions, or that surround the transcription start sites of silent genes, show large variation in positioning across different cells but are highly uniformly spaced along the nucleosome array; and second, nucleosomes that surround the transcription start sites of active genes and DNase I hypersensitive sites show little variation in positioning across different cells but are relatively heterogeneously spaced along the nucleosome array. We found a bimodal distribution of nucleosome spacing at DNase I hypersensitive sites, which corresponds to inaccessible and accessible states and is associated with nucleosome variation and variation in accessibility across cells. Nucleosome variation is smaller within single cells than across cells, and smaller within the same cell type than across cell types. A large fraction of naive CD4 T cells and mouse embryonic stem cells shows depleted nucleosome occupancy at the de novo enhancers detected in their respective differentiated lineages, revealing the existence of cells primed for differentiation to specific lineages in undifferentiated cell populations.

Dataset Information

Aberrant nucleosome organization in mouse SCNT embryos revealed by ULI-MNase-seq.

Publications

Aberrant nucleosome organization in mouse SCNT embryos revealed by ULI-MNase-seq.

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