Zscan10 is dispensable for maintenance of pluripotency in mouse embryonic stem cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE76143 | GEO | 2015/12/20
SECONDARY ACCESSION(S): PRJNA306423
REPOSITORIES: GEO
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Project description:Zinc finger and SCAN domain-containing 10 (Zscan10, also known as Zfp206) encodes a transcription factor that has been reported to be involved in the maintenance of pluripotency in mouse embryonic stem (ES) cells. Here we generated inducible knockout ES cells for Zscan10 using the Cre-loxP system and analyzed its function. We succeeded in establishing Zscan10-null ES cells and confirmed their pluripotency by the generation of chimeric embryos. Our results clearly indicate that Zscan10 is dispensable for the ability of self-renewal and differentiation in ES cells. An inducible knockout ES cell line of Zscan10 with the Cre-loxP system was generated and gene expression was measured without Zscan10 deletion or constitutive knocked out cells. Each sample was prepared in triplicate.
2015-12-20 | E-GEOD-76143 | biostudies-arrayexpress
Project description:We showed the function of Rex1 in mouse ES cells as well as in embryos using the conventional gene targeting strategy. Our results clearly indicated that Rex1 function is dispensable for both the maintenance of pluripotency in ES cells and the development of embryos. However, Rex1-/- ES cells showed the defect to induce a subset of the marker genes of visceral endoderm, when differentiated as embryoid body, as found in EC cells. Conclusion: Rex1 should be regarded just as a marker of pluripotency without functional significance like the activity of alkaline phosphatase. Keywords: genetic modification design,reference design,replicate design
2008-09-13 | GSE12753 | GEO
Project description:Zinc finger and SCAN domain containing 10 (Zscan10) was identified as a novel transcription factor in osteoclast differentiation by our previous study. However, the biological functions of Zscan10 have not been fully understood except its role in maintenance of genome stability and pluripotency in embryonic stem cells. Therefore, the purpose of this study is clarification of Zscan10 function in somatic cells, especially during osteoclast differentiation. First, Zscan10 KO RAW264 (KO) cells were established using CRISPR/Cas9 system and single cell sorting. Then, Control (Ctrl) and KO cells were differentiated into osteoclast by RANKL stimulation. As a result, TRAP activity and expression levels of differentiation marker genes, such as Nfatc1, were significantly increased and the expression of inhibitory factors, such as Irf8, was decreased in KO cells as compared to Ctrl cells. These results suggested that Zscan10 might regulate transcription of the genes which negatively control osteoclastogenesis. To understand the transcriptomes controlled by Zscan10, RNA-seq was performed and the stringent analyses identified significantly down-regulated Haptoglobin (Hp) in KO cells. Additionally, Zscan10 binding sequence was located near the genomic region of Hp gene locus. ChIP against Zscan10 followed by qPCR for the region revealed that Zscan10 binds to the region located near Hp gene locus and transcript start site, suggesting that Zscan10 may regulate transcription of Hp. Next, to examine the effect of Hp under Zscan10 mediated osteoclastogenesis, KO cells were treated with recombinant Hp (rHp). As a result, Hp treatment could suppress the elevated TRAP activity of KO cells without affecting cell viability. Furthermore, Hp KO mice exhibit decreased bone mass and increased osteoclast number, and Hp has been reported to be involved in suppression of osteoclastogenesis. Also, In hemolytic disease, Hp had been decreased and also bone density. These facts suggested that Zscan10 negatively regulates osteoclast differentiation through transcription of Hp.
2019-03-25 | GSE121320 | GEO
Project description:The transcription factor Oct3/4 is essential to maintain pluripotency in mouse embryonic stem (ES) cells. It was reported that the Xpc DNA repair complex is involved in this process. Here we examined the role of Xpc on the transcriptional activation of the target genes by Oct3/4 using the inducible knockout strategy. We found that the removal of the C-terminal region of Xpc, including the interaction sites with Rad23 and Cetn2, showed faint impact on the gene expression profile of ES cells and the functional Xpc-ΔC ES cell lines retained proper gene expression profile as well as pluripotency to contribute chimeric embryos. These data indicated that the C-terminal region of Xpc is dispensable for the transcriptional activity of Oct3/4 in mouse ES cells. An inducible knockout ES cell line of Xpc with the Cre-loxP system was generated and gene expression was measured without Xpc deletion, after 4 days of Xpc deletion or constitutive knocked out cells. Each sample was prepared in triplicate.
2015-06-09 | E-GEOD-69640 | biostudies-arrayexpress
Project description:Abstract: The Promyelocytic Leukemia protein (PML) and its associated nuclear bodies have re-cently emerged as essential factors for maintaining the characteristics of embryonic stem (ES) cells. However, the full repertoire of PML driven gene regulatory events in ES cells is not resolved. In this report we have studied the role of PML in shaping the proteomic and SUMO proteomic landscape in ES cells. Our analysis of the PML KD proteome revealed a suppression of proteins related with self-renewal and an up-regulation of proteins vital for translation and proteasome functions, reflecting a cellular transition from pluripotency to differentiation. Major targets of PML-directed sumoylation include pluripotency factors, chromatin organizers and cell cycle regulators. We demonstrate that PML promotes the sumoylation of SALL1 and CDCA8, two proteins that are highly expressed in undifferentiated ES cells. SALL1 sumoylation increases the activation of the Wnt pathway, contributing to its ability to inhibit ES cell differentiation. Similarly, CDCA8 sumoylation enhances its capacity to promote cell proliferation. Our results demonstrate that PML maintains ES cell functions by modulating the abundance or sumoylation of key regulators involved in pluripotency and cell cycle progression.
2025-05-07 | PXD058024 | Pride
Project description:Abstract: The Promyelocytic Leukemia protein (PML) and its associated nuclear bodies have re-cently emerged as essential factors for maintaining the characteristics of embryonic stem (ES) cells. However, the full repertoire of PML driven gene regulatory events in ES cells is not resolved. In this report we have studied the role of PML in shaping the proteomic and SUMO proteomic landscape in ES cells. Our analysis of the PML KD proteome revealed a suppression of proteins related with self-renewal and an up-regulation of proteins vital for translation and proteasome functions, reflecting a cellular transition from pluripotency to differentiation. Major targets of PML-directed sumoylation include pluripotency factors, chromatin organizers and cell cycle regulators. We demonstrate that PML promotes the sumoylation of SALL1 and CDCA8, two proteins that are highly expressed in undifferentiated ES cells. SALL1 sumoylation increases the activation of the Wnt pathway, contributing to its ability to inhibit ES cell differentiation. Similarly, CDCA8 sumoylation enhances its capacity to promote cell proliferation. Our results demonstrate that PML maintains ES cell functions by modulating the abundance or sumoylation of key regulators involved in pluripotency and cell cycle progression.
2025-05-23 | PXD057990 | Pride
Project description:Oct4 is an essential regulator of embryonic stem (ES) cell pluripotency in vivo and in vitro, as well as a key mediator of the reprogramming of somatic cells to form induced pluriopotent stem (iPS) cells. It is not known whether activation and/or repression of specific genes by Oct4 is relevant to these functions. Here we show that fusion proteins containing the coding sequence of Oct4 or Xlpou91 (the Xenopus homologue of Oct4) fused to activating regions, but not those fused to repressing regions, behave as Oct4, suppressing differentiation and promoting maintenance of undifferentiated phenotypes in vivo and in vitro. An Oct4 activation domain fusion supported ES cell self-renewal in vitro at lower concentrations than required for Oct4 while alleviating the ordinary requirement for the cytokine LIF. At still lower levels of the fusion, LIF-dependence was restored. We conclude that the necessary and sufficient function of Oct4 in promoting pluripotency is to activate specific target genes. Two independent clones from each cell line were used as biological replications. Cy3-CTP-labelled sample targets were prepared from 2.5 ?g of total RNA using the Low RNA Input Fluorescent Linear Amplification Kit (Agilent). Cy5-CTP-labelled reference target was produced from 2.5 ?g of Stratagene Universal Mouse Reference RNA. Purified target RNA were hybridized to the NIA Mouse 44K Microarray v3.0 (whole genome 60-mer oligo arrays, Agilent Technology, design ID 015087) (Carter et al., 2005) according to the manufacturer's protocol (Two-Colour Microarray-Based Gene Expression Analysis Protocol, Version 5.0.1). Slides were scanned with an Agilent DNA Microarray Scanner (model G2505-64120) at 100% PMT for the Cy3 channel and 10% PMT for the Cy5 channel with a scan resolution of 5µm. Data was analyzed using NIA Array Analysis (Sharov et al., 2005) using standard statistical conditions (FDR<0.05, 2-fold expression levels change) to unveil genes with changes in expression levels between the samples and controls.
2012-02-27 | E-GEOD-24563 | biostudies-arrayexpress
Project description:Many lincRNAs have been identified as essential regulators for ESC pluripotency through siRNA-based screening. However, the biological and molecular functions of the large intergenic noncoding RNAs (lincRNAs) remain unclear. Here, we employed CRISPR/Cas9-mediated knockout technology to explore the functions of 8 previously annotated lincRNAs in ESCs. Unexpectedly, it was found that most lincRNAs are dispensable for pluripotency maintenance, cloning formation ability, proliferation and apoptosis in ESCs when we disrupted these lincRNAs individually or in combinations. Single-cell transcriptomic analysis of a set of lncRNA knockout ESCs showed consistent results, indicating that previously observed functions of lincRNAs on ESC pluripotency maintenance may be due to off-targeted effects. Interestingly, deletion and transcriptional repression of linc1343 disrupts embryoid body (EB) differentiation in ESC. Notably, GM17300 and two small RNAs (Snora73a, Snora73b) derived from the lnc1343 locus are highly expressed in ESCs. Further overexpression of Snora73a and Snora73b in linc1343 knockout cells rescue the phenotype, suggesting that snoRNAs derived from linc1343 are the responders for EB differentiation. Taken together, our results demonstrate the divergent effects of lincRNAs on regulating ESC pluripotency and EB differentiation.
2024-08-07 | GSE225238 | GEO
Project description:Many lincRNAs have been identified as essential regulators for ESC pluripotency through siRNA-based screening. However, the biological and molecular functions of the large intergenic noncoding RNAs (lincRNAs) remain unclear. Here, we employed CRISPR/Cas9-mediated knockout technology to explore the functions of 8 previously annotated lincRNAs in ESCs. Unexpectedly, it was found that most lincRNAs are dispensable for pluripotency maintenance, cloning formation ability, proliferation and apoptosis in ESCs when we disrupted these lincRNAs individually or in combinations. Single-cell transcriptomic analysis of a set of lncRNA knockout ESCs showed consistent results, indicating that previously observed functions of lincRNAs on ESC pluripotency maintenance may be due to off-targeted effects. Interestingly, deletion and transcriptional repression of linc1343 disrupts embryoid body (EB) differentiation in ESC. Notably, GM17300 and two small RNAs (Snora73a, Snora73b) derived from the lnc1343 locus are highly expressed in ESCs. Further overexpression of Snora73a and Snora73b in linc1343 knockout cells rescue the phenotype, suggesting that snoRNAs derived from linc1343 are the responders for EB differentiation. Taken together, our results demonstrate the divergent effects of lincRNAs on regulating ESC pluripotency and EB differentiation.
2024-08-07 | GSE225237 | GEO
Project description:Mouse embryonic stem (ES) cells are locked into self-renewal by shielding from inductive cues. Release from this ground state in minimal conditions offers a system for delineating developmental progression from naive pluripotency. Here we examined the initial transition process. The ES cell population behaves asynchronously. We therefore exploited a short-half-life Rex1::GFP reporter to isolate cells either side of exit from naive status. Differentiation of Rex1-GFPd2 ES cells was initiated by withdrawing 2i (Kalkan et al., 2016). Undifferentiated 2i-cells and post-2i withdrawal differentiating populations (16h, 25h-Rex1-High, 25h-Rex1-Low) were subjected to proteomic analysis by Mass Spectrometry.
2017-02-22 | PXD005581 | Pride