Journal: PLoS ONE

Article Title: Brg1 Is Required for Cdx2-Mediated Repression of Oct4 Expression in Mouse Blastocysts

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: (A) qRT-PCR analysis of Brg1, Cdx2, and Oct4 transcripts in Brg1 KD 8-cell embryos, morulae, and blastocysts. Data were normalized to Ubtf (house keeping gene) and are relative to control embryos at each stage; black line = 1. Asterisk denotes significant difference between Brg1 KD and control blastocysts (p<0.05). (B) ICC analysis of Oct4 and Cdx2 expression in Brg1 KD 8-cell embryos, morulae, and blastocysts. Nuclei were counter stained with DAPI (blue). (C) Brg1 represses Oct4 expression in a dose dependent manner. One-cell embryos were injected with 0 µM (control), 0.1 µM, 1 µM, or 100 µM Brg1 siRNA and cultured to the blastocyst stage. Real-time qPCR was used to analyze the levels of Brg1 and Oct4 transcripts. Data were normalized to Ubtf and are relative to control blastocysts; dashed line = 1.

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques: Quantitative RT-PCR, Control, Expressing, Staining, Injection, Cell Culture

Journal: PLoS ONE

Article Title: Brg1 Is Required for Cdx2-Mediated Repression of Oct4 Expression in Mouse Blastocysts

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: (A) ICC analysis of Oct4 and Cdx2 in Brg1 KD and control blastocysts. In control blastocysts (a–d) Oct4 expression (green) is restricted to the ICM and is largely absent in the Cdx2-positive (red) trophectoderm. In contrast, in Brg1 KD blastocysts (e–h) Oct4 is widely expressed in both the ICM and cdx2-positive (yellow) trophectoderm. Arrowheads denote co-expression of Oct4 and Cdx2. Nuclei were counter stained with DAPI (blue). (B) Quantification of the average number of cells in Brg1 KD and control blastocysts expressing Oct4, Cdx2, and Oct4 & Cdx2 (double expression). Asterisks denote statistical significance (p<0.05) between Brg1 KD and control blastocysts. A total of 25 Brg1 KD blastocysts and 15 control blastocysts were analyzed.

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques: Control, Expressing, Staining

Journal: PLoS ONE

Article Title: Brg1 Is Required for Cdx2-Mediated Repression of Oct4 Expression in Mouse Blastocysts

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: (A) Combined depletion of Brg1 and Cdx2 augments Oct4 expression in blastocysts. qRT-PCR analysis of Oct4 transcripts in Brg1 KD blastocysts, Cdx2 KD blastocysts, and Brg1 & Cdx2 double KD blastocysts. Data were normalized to Ubtf (house keeping gene) and are relative to control blastocysts; dashed line = 1. Different letters denote statistical significance in Oct4 transcripts (p<0.05). These experiments were replicated using a total of 5 biological replicates. (B) Co-immunoprecipitation and western blot analysis of Brg1 and Cdx2 in TS cells. Brg1 was immunoprecipitated using a rabbit anti-serum. Recovery of Cdx2 was measured by western blot analysis. Cdx2 is enriched in the Brg1 IP samples and not in the control IgG samples. This assay was repeated a total of 4 times using different batches of TS cells. (C) Confocal immunofluorescence analysis of Brg1 and Cdx2 in blastocysts. Co-localization of endogenous Brg1 and Cdx2 in trophectoderm nuclei was determined using specific antibodies for Brg1 and Cdx2. Nuclei were counterstained with DAPI. White box represents magnified region in bottom panel. Arrow denotes blastocyst ICM. (D) Confirmation of Flag-Cdx2 expression in induced ES cells. Western blot and immunofluorescence analysis of Flag-Cdx2 expression at 24 hours following removal of doxycycline. (E) ChIP analysis of Brg1 and Cdx2 binding to the Oct4 promoter in Cdx2-inducible ES cells. qRT-PCR was used to determine the relative enrichment of Brg1 and Flag-Cdx2 at the Oct4 ARE versus an intergenic region in uninduced and induced ES cell extracts. A non-specific rabbit IgG was included as a negative control. Data were normalized to 1% input DNA. Asterisks denote significant differences between uninduced and induced samples (p<0.05). These experiments were replicated 3 to 4 times using two different batches of Cdx2-inducible ES cell extracts.

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques: Expressing, Quantitative RT-PCR, Control, Immunoprecipitation, Western Blot, Immunofluorescence, Binding Assay, Negative Control

Journal: PLoS ONE

Article Title: Brg1 Is Required for Cdx2-Mediated Repression of Oct4 Expression in Mouse Blastocysts

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: Schematic diagram of Oct4 regulation in blastocysts. In the trophectoderm Brg1 is recruited to the ARE of the Oct4 promoter via Cdx2. Once at the Oct4 promoter Brg1 and Cdx2 facilitate recruitment of additional co-repressors to repress transcription.

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques:

Journal: Cell

Article Title: Nervous System Regionalization Entails Axial Allocation before Neural Differentiation

doi: 10.1016/j.cell.2018.09.040

Figure Lengend Snippet:

Article Snippet: Mouse monoclonal anti-CDX2 , Abcam , 157524; RRID: AB_2721036.

Techniques: Recombinant, Cloning, RNA In Situ Hybridization, Software

Journal: Human molecular genetics

Article Title: ELF5-enforced transcriptional networks define an epigenetically regulated trophoblast stem cell compartment in the human placenta.

doi: 10.1093/hmg/ddq128

Figure Lengend Snippet: Figure 1. Genomic organization of the human ELF5 locus and transcript isoform expression in placenta and trophoblast cell lines. (A) Diagram of the exon–intron structure of the human ELF5 locus and annotated splice var- iants. Position of primers used is indicated. Filled boxes represent open- reading frames and open boxes represent untranslated regions. (B) RT–PCR analysis with isoform-specific and common primers reveals that ELF5-2b is the expressed splice variant in placenta and the trophoblast-like cell line TCL-1, but that it is absent from the first trimester mesenchymal-like cell line TCL-2. (C) RT–PCR with primers spanning exons 3 and 4 demonstrates that the annotated ELF5-2bDex3/4 variant is not present in placenta and chor- iocarcinoma and trophoblast-like cell lines JEG-3 and TCL-1.

Article Snippet: For each immunoprecipitation reaction, 50 mg of chromatin was pre-cleared and incubated overnight at 48C with 5 mg of anti-ELF5 antibody (Santa Cruz Biotechnology), anti-CDX2 antibody (BioGenex) or control antibody bound to Protein G sepharose beads (Amersham).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Variant Assay

Journal: Human molecular genetics

Article Title: ELF5-enforced transcriptional networks define an epigenetically regulated trophoblast stem cell compartment in the human placenta.

doi: 10.1093/hmg/ddq128

Figure Lengend Snippet: Figure 2. Expression of trophoblast stem cell genes and epigenetic regulation of ELF5 in placenta throughout gestation. (A) RT–PCR analysis of ELF5, CDX2 and EOMES (i.e. genes important for trophoblast stem cell self-renewal and proliferation in the mouse) on human placental villous samples ranging from 7 weeks of gestation to term. Four independent term placental samples were investigated. The choriocarcinoma cell line JEG-3 was included as control. Colour-inverted photographs of ethidium bromide stained gels are shown. All three genes are expressed in placenta, but CDX2 is not detected from the second trimester onwards even when the PCRs are over-cycled. (B) Quantitative RT–PCR (qPCR) analysis of ELF5, CDX2 and EOMES on the same samples used in (A). ELF5 is down- regulated in second and third trimesters, whereas no overall regulation with gestational age was observed for EOMES. (C) Comparison of expression levels between first trimester and term. ELF5 expression is significantly reduced at term when compared with first trimester, CDX2 is absent from term placentas. (D) Bisulphite sequencing analysis of the ELF5 promoter region. Filled circles indicate methylated cytosine residues. ELF5 is extremely hypomethylated in the first trimester and acquires higher DNA methylation levels in second and third trimester, correlating with transcriptional down-regulation at these stages. (E) DNA methylation analysis of an extended region between 2400 bp and +400 bp around the transcriptional start site of ELF5. Hypomethylation correlates with ELF5 expression in JEG-3 cells and, conversely, ELF5 is hypermethylated and not expressed in TCL-2 cells. The methylation pattern in TCL-1 cells reveals a critical stretch of five CpG residues (grey box) at the immediate transcriptional start site that needs to be unmethylated for ELF5 to be expressed.

Article Snippet: For each immunoprecipitation reaction, 50 mg of chromatin was pre-cleared and incubated overnight at 48C with 5 mg of anti-ELF5 antibody (Santa Cruz Biotechnology), anti-CDX2 antibody (BioGenex) or control antibody bound to Protein G sepharose beads (Amersham).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Control, Staining, Quantitative RT-PCR, Comparison, Bisulfite Sequencing, Methylation, DNA Methylation Assay

Journal: Human molecular genetics

Article Title: ELF5-enforced transcriptional networks define an epigenetically regulated trophoblast stem cell compartment in the human placenta.

doi: 10.1093/hmg/ddq128

Figure Lengend Snippet: Figure 3. Immunofluorescence localization of ELF5 to cytotrophoblasts in the human placenta. (A) Overview of 11 week placental villous cross-section shows ELF5 localization to nuclei of villous cytotrophoblasts, but absence from nuclei of the overlying syncytiotrophoblast layer. Cytotrophoblasts are a proliferative cell population that continuously divide to replenish the overlying syncytium. (B) Co-localization with cytokeratin 7 (CK7) confirms the trophoblast identity of ELF5-positive cells. (C) Confocal image of a double staining of ELF5 and the villous cytotrophoblast marker SPINT1 (also known as HAI-1) shows that every ELF5-positive nucleus resides within the cytotrophoblast layer. Top row 6 week, bottom row 11 week placenta. (D) Confocal image analysis of an 11 week villous section stained for ELF5 and the extravillous cytotrophoblast (EVT) marker integrin alpha-5 (ITGA5). ELF5 is detected only in nuclei at the proliferative base, but not further distal along the EVT column where cells adopt an invasive phenotype and lose proliferative potential. (E) ELF5 is also absent from post- mitotic interstitial and endovascular EVTs within the decidual bed.

Article Snippet: For each immunoprecipitation reaction, 50 mg of chromatin was pre-cleared and incubated overnight at 48C with 5 mg of anti-ELF5 antibody (Santa Cruz Biotechnology), anti-CDX2 antibody (BioGenex) or control antibody bound to Protein G sepharose beads (Amersham).

Techniques: Double Staining, Marker, Staining

Journal: Human molecular genetics

Article Title: ELF5-enforced transcriptional networks define an epigenetically regulated trophoblast stem cell compartment in the human placenta.

doi: 10.1093/hmg/ddq128

Figure Lengend Snippet: Figure 4. CDX2 identifies a subset of ELF5-positive cytotrophoblasts as a TS-like compartment that is regulated by FGFR2. (A) ELF5 co-localizes with FGFR2 in villous cytotrophoblasts as identified by confocal image analysis of double immunofluorescence stainings of 11 week placental sections. Since FGF signalling has been implicated in TS cell proliferation in mice and humans and can activate ELF5 expression in other tissues, FGF/FGFR2 may induce ELF5 expression within a putative TS cell niche in the human placenta. (B) Double staining of a 6 week placental section for ELF5 and CDX2. Larger groups of CDX2-positive cells are detected only in early gestation up to 8.5–9 weeks. CDX2 is mostly co-expressed with ELF5 (arrowheads). (C) Dual labelling of 6 week placental section for CDX2 and the proliferation marker Ki67. CDX2-expressing cytotrophoblasts preferentially stain positive for Ki67, indicating their high proliferation rate. CDX2 and Ki67 are restricted to the proximal end of cytotrophoblast cell columns (highlighted by the boxed area). The white arrows indicate the direction of progressive extravillous trophoblast (EVT) differentiation and migration.

Article Snippet: For each immunoprecipitation reaction, 50 mg of chromatin was pre-cleared and incubated overnight at 48C with 5 mg of anti-ELF5 antibody (Santa Cruz Biotechnology), anti-CDX2 antibody (BioGenex) or control antibody bound to Protein G sepharose beads (Amersham).

Techniques: Expressing, Double Staining, Marker, Staining, Migration

Journal: Human molecular genetics

Article Title: ELF5-enforced transcriptional networks define an epigenetically regulated trophoblast stem cell compartment in the human placenta.

doi: 10.1093/hmg/ddq128

Figure Lengend Snippet: Figure 5. Inter-regulatory network of trophoblast transcription factors CDX2, EOMES and ELF5. (A) Chromatin immunoprecipitation assays show that CDX2 binds to the ELF5 promoter region in JEG-3 and TCL-1 cells where ELF5 is hypomethylated and expressed, but not in TCL-2 cells where ELF5 is hypermethylated and repressed. (B) In turn, ELF5 binds to the CDX2 and EOMES promoter regions in JEG-3 and TCL-1 cells where it is expressed, but not in TCL-2 cells from which it is absent, thereby establishing a transcrip- tional feedback loop between all three transcription factors. Binding to the EOMES promoter region was more consistent and is indicative of a more effi- cient, stronger interaction than with the CDX2 upstream region, consistent with results observed in mouse trophoblast (9).

Article Snippet: For each immunoprecipitation reaction, 50 mg of chromatin was pre-cleared and incubated overnight at 48C with 5 mg of anti-ELF5 antibody (Santa Cruz Biotechnology), anti-CDX2 antibody (BioGenex) or control antibody bound to Protein G sepharose beads (Amersham).

Techniques: Chromatin Immunoprecipitation, Binding Assay

Journal: Human molecular genetics

Article Title: ELF5-enforced transcriptional networks define an epigenetically regulated trophoblast stem cell compartment in the human placenta.

doi: 10.1093/hmg/ddq128

Figure Lengend Snippet: Figure 6. Trophoblast transcription factor expression and epigenetic regulation of ELF5 in human ES cells and derived trophoblast cell lines. (A) Initial bisul- phite sequencing analysis of two pooled hES cell lines and derived trophoblast cells indicates a high degree of DNA methylation at the ELF5 promoter despite the limited trophoblast differentiation potential. (B) RT–PCR and (C) qPCR analysis for trophoblast transcription factors ELF5, CDX2 and EOMES on six differ- ent hES cells lines (Shef1, Shef4–7, H7), including one subclone with an abnormal karyotype (Shef5a), two derived cytotrophoblast cell lines (TrophH7 and TrophShef4), the JEG-3, TCL-1 and TCL-2 cell lines, an 8+4 week placenta for relative comparison of expression levels and a colorectal cancer cell line (DKO4) as positive control for CDX2 expression (27). Colour-inverted photographs of ethidium bromide stained gels are shown. ELF5 is detectable in some hES cell lines, albeit at very low levels. Higher expression levels of CDX2 and EOMES may relate to their function within the embryonic lineage and is not directly indicative of trophoblast differentiation potential. Strikingly, in contrast to their expression in placenta, all three genes are absent from the hES-derived tropho- blast cell lines. (D) Normalization of qPCR data to Shef6, one of the most highly ELF5 expressing hES cell lines, in comparison with JEG-3, TCL-1 and TCL-2 cell lines as well as a first trimester placenta sample demonstrates the comparatively negligible amount of ELF5 expression in hES cells that is approximately 300-fold less than in normal trophoblast in vivo. (E) Bisulphite sequencing analysis of the ELF5 promoter in three different hES cell lines and two derived trophoblast cell lines shows relatively little epigenetic variability between different hES cell lines. Hypermethylation correlates with extremely low ELF5 expression levels. (F) Elf5 is also highly methylated in three independent mouse epiblast stem cell lines and (G) in two human-induced pluripotent stem cell lines derived from kereatinocytes and fibroblasts.

Article Snippet: For each immunoprecipitation reaction, 50 mg of chromatin was pre-cleared and incubated overnight at 48C with 5 mg of anti-ELF5 antibody (Santa Cruz Biotechnology), anti-CDX2 antibody (BioGenex) or control antibody bound to Protein G sepharose beads (Amersham).

Techniques: Expressing, Derivative Assay, Sequencing, DNA Methylation Assay, Reverse Transcription Polymerase Chain Reaction, Comparison, Positive Control, Staining, In Vivo, Bisulfite Sequencing, Methylation

Journal: Advanced Science

Article Title: Capture of Totipotency in Mouse Embryonic Stem Cells in the Absence of Pdzk1

doi: 10.1002/advs.202408852

Figure Lengend Snippet: Extended potency of Pdzk1 ‐KO ESCs. A) Sense (red) and antisense (green) insertions of PB in the Pdzk1 gene body from a previous screen of iTSCs via a haploid system. The rectangles indicate the exons. B) Schematic diagram of the strategy for deleting Pdzk1 in WT‐ESCs via the CRISPR/Cas9 system. C) Genotype PCR results of Pdzk1 ‐KO subclones. D) Genotyping of Pdzk1 ‐KO ESCs via DNA sequencing. E) Morphological images of the induced cell cultures of Pdzk1 ‐KO ESCs and WT‐ESCs before and after cell sorting with a TSC‐specific antibody (CDCP1). Scale bar, 100 µm. F) Percentages of CDCP1+ cells in induced cultures of Pdzk1 ‐KO ESCs and WT‐ESCs. G) Morphological images of Pdzk1 ‐KO iTSCs and WT‐TSCs. Scale bar, 100 µm. H) Immunofluorescence of TSC‐specific markers (CDX2 and EOMES) in Pdzk1 ‐KO iTSCs and WT‐TSCs. DNA was stained with Hoechst 33342. Scale bar, 50 µm. I) Morphological images of Pdzk1 ‐KO iXENs and WT‐ESC‐derived iXENs. Scale bar, 100 µm. J) Immunofluorescence of the XEN‐specific marker (GATA6) in Pdzk1 ‐KO iXENs and WT‐ESC iXENs. DNA was stained with Hoechst 33342. Scale bar, 50 µm. K) Summary of the potencies of Pdzk1 ‐KO ESCs and WT‐ESCs.

Article Snippet: The primary antibodies used were CDX2 (BioGenes, MU392A), OCT4 (Abcam, ab181557), GATA6 (CST, 5851), EOMES (Abcam, ab23345), TFAP2C (Santa Cruz, sc‐12762), and YAP1 (ABclonal, A1002).

Techniques: CRISPR, DNA Sequencing, FACS, Immunofluorescence, Staining, Derivative Assay, Marker

Journal: Advanced Science

Article Title: Capture of Totipotency in Mouse Embryonic Stem Cells in the Absence of Pdzk1

doi: 10.1002/advs.202408852

Figure Lengend Snippet: Advanced developmental potential of Pdzk1 ‐KO ESCs in chimeras. A) Schematic overview of the developmental contribution of microinjection of Pdzk1 ‐KO ESCs and WT‐ESCs into chimeric embryos. B) Immunofluorescence of OCT4 (red) in chimeric blastocysts derived from GFP‐labeled (green) Pdzk1 ‐KO ESCs and WT‐ESCs. DNA was stained with Hoechst 33342 (blue). Scale bar, 50 µm. C) Immunofluorescence of CDX2 (red) in chimeric blastocysts derived from GFP‐labeled (green) Pdzk1 ‐KO ESCs and WT‐ESCs. DNA was stained with Hoechst 33342 (blue). Scale bar, 50 µm. D) Immunofluorescence of GATA6 (red) in chimeric blastocysts derived from GFP‐labeled (green) Pdzk1 ‐KO ESCs and WT‐ESCs. DNA was stained with Hoechst 33342 (blue). Scale bar, 50 µm. E) Images of E12.5 chimeras (including fetus, placenta and yolk sac) derived from GFP‐labeled Pdzk1 ‐KO ESCs, with GFP‐labeled WT‐ESC‐derived chimeras used as a control. The white arrowheads indicate the fetus, placenta and yolk sac from the chimeric embryo. Scale bar, 5 mm. F) Heatmap representation of genes predominantly expressed in GFP+ cells from the chimeric placenta and chimeric yolk sac. The WT‐placenta and WT‐yolk sac data were downloaded from a previous study (Andergassen et al., 2017, Elife). Representative lineage markers are indicated.

Article Snippet: The primary antibodies used were CDX2 (BioGenes, MU392A), OCT4 (Abcam, ab181557), GATA6 (CST, 5851), EOMES (Abcam, ab23345), TFAP2C (Santa Cruz, sc‐12762), and YAP1 (ABclonal, A1002).

Techniques: Microinjection, Immunofluorescence, Derivative Assay, Labeling, Staining, Control

Journal: Advanced Science

Article Title: Capture of Totipotency in Mouse Embryonic Stem Cells in the Absence of Pdzk1

doi: 10.1002/advs.202408852

Figure Lengend Snippet: Generation and development of Pdzk1 ‐KO ESC‐derived blastoids. A) Blastoid formation efficiencies of Pdzk1 ‐KO ESCs and Dyrk1a ‐KO ESCs, with WT‐ESCs serving as a negative control. B) Images of Pdzk1 ‐KO blastoids, Dyrk1a ‐KO blastoids and E4.5 mouse blastocysts. Scale bar, 100 µm. C) Total number of cells in each E4.5 mouse blastocyst (n = 5) and each blastoid ( Pdzk1 ‐KO, n = 15; Dyrk1a ‐KO, n = 19). D) The diameter of each E4.5 mouse blastocyst (n = 19) and each blastoid ( Pdzk1 ‐KO, n = 19; Dyrk1a ‐KO, n = 19) separately. E) Immunofluorescence of OCT4 (red) and CDX2 (green) in Pdzk1 ‐KO blastoids. WT blastocysts were used as controls. DNA was stained with Hoechst 33342 (blue). Scale bar, 50 µm. F) Immunofluorescence of GATA6 (red) and CDX2 (green) in Pdzk1 ‐KO blastoids. WT blastocysts were used as controls. DNA was stained with Hoechst 33342 (blue). Scale bar, 50 µm. G) Schematic overview of the establishment of BESCs, BTSCs, and BXENs derived from GFP‐labeled blastoids. H) GFP and immunofluorescence images of BESCs, BTSCs, and BXENs derived from Pdzk1 ‐KO blastoids with specific markers. Scale bar, GFP images, 100 µm; Immunofluorescence images, 50 µm. I) Images of in vitro cultured (IVC) blastoids on Days 1–4. Scale bars, 100 µm. J) Immunofluorescence of OCT4 and CDX2 in IVC blastoids on Day 4. Scale bar, 50 µm. K) Immunofluorescence of CDX2 and GATA6 in IVC blastoids on Day 4. Scale bar, 50 µm.

Article Snippet: The primary antibodies used were CDX2 (BioGenes, MU392A), OCT4 (Abcam, ab181557), GATA6 (CST, 5851), EOMES (Abcam, ab23345), TFAP2C (Santa Cruz, sc‐12762), and YAP1 (ABclonal, A1002).

Techniques: Derivative Assay, Negative Control, Immunofluorescence, Staining, Labeling, In Vitro, Cell Culture

Journal: Advanced Science

Article Title: Capture of Totipotency in Mouse Embryonic Stem Cells in the Absence of Pdzk1

doi: 10.1002/advs.202408852

Figure Lengend Snippet: Single‐cell transcriptome analysis of Pdzk1 ‐KO blastoids and Pdzk1 ‐KO ESCs. A) UMAP plots of Pdzk1 ‐KO blastoids and WT blastocysts. B) Comparison of cells derived from Pdzk1 ‐KO blastoids and WT blastocysts. This panel shows lineage assignments: epiblast (purple), primitive endoderm (green) and trophectoderm (blue). C) The expression of representative marker genes for epiblast ( Oct4 ), primitive endoderm ( Gata6 ) and trophectoderm ( Cdx2 ) in Pdzk1 ‐KO blastoids at the single‐cell level. D) Principal component analysis (PCA) of Pdzk1 ‐KO blastoids compared with published datasets of mouse embryos. The embryonic stages are indicated. E) PCA of Pdzk1 ‐KO ESCs, WT‐ESCs, ciTotiSCs, TBLCs, L. EPSCs, D. EPSCs, TBLCs and mouse embryos at different stages at the single‐cell level. F) Hierarchical clustering analysis of Pdzk1 ‐KO ESCs, WT‐ESCs, ciTotiSCs, TBLCs, L. EPSCs, D. EPSCs, TBLCs and mouse embryos at different stages at the single‐cell level.

Article Snippet: The primary antibodies used were CDX2 (BioGenes, MU392A), OCT4 (Abcam, ab181557), GATA6 (CST, 5851), EOMES (Abcam, ab23345), TFAP2C (Santa Cruz, sc‐12762), and YAP1 (ABclonal, A1002).

Techniques: Comparison, Derivative Assay, Expressing, Marker

Journal: Advanced Science

Article Title: Capture of Totipotency in Mouse Embryonic Stem Cells in the Absence of Pdzk1

doi: 10.1002/advs.202408852

Figure Lengend Snippet: Validation of lineage inducers in the totipotent state. A) Expression levels of OE genes ( C1qa , C1qb , Fgf5 , and Cdx2 ) in OE‐ESCs, with the empty vector group and WT‐ESCs used as controls. The data are presented as the means ± SDs. *** p < 0.001. B) Images of E12.5 chimeras (including fetuses, placentas and yolk sacs) derived from GFP‐labeled C1qa ‐OE, C1qb ‐OE, Fgf5 ‐OE, and Cdx2 ‐OE ESCs, with the GFP‐labeled empty vector group used as a control. Scale bar, 5 mm. C) Summary of C1qa ‐OE, C1qb ‐OE, Fgf5 ‐OE, and Cdx2 ‐OE ESCs contributing to three lineages according to chimeric assays. D) Volcano plot of DEGs between Pdzk1 ‐KO ESCs and WT‐ESCs. C1qa , C1qb , Fgf5 , and Cdx2 are highlighted.

Article Snippet: The primary antibodies used were CDX2 (BioGenes, MU392A), OCT4 (Abcam, ab181557), GATA6 (CST, 5851), EOMES (Abcam, ab23345), TFAP2C (Santa Cruz, sc‐12762), and YAP1 (ABclonal, A1002).

Techniques: Biomarker Discovery, Expressing, Plasmid Preparation, Derivative Assay, Labeling, Control

Journal: Advanced Science

Article Title: Capture of Totipotency in Mouse Embryonic Stem Cells in the Absence of Pdzk1

doi: 10.1002/advs.202408852

Figure Lengend Snippet: Single‐cell transcriptome profiles of Pdzk1 ‐KO ESCs and WT‐ESCs. A) UMAP plots of Pdzk1 ‐KO ESCs and WT‐ESCs. B) UMAP plots of Pdzk1 ‐KO ESCs, WT‐ESCs, WT‐TSCs, and WT‐XENs. C) Normalized expression of C1qa , C1qb , Fgf5 , and Cdx2 in Pdzk1 ‐KO ESCs and WT‐ESCs according to the UMAP at the single‐cell level. D) Dot plots indicating the expression of C1qa , C1qb , Fgf5 , and Cdx2 in Pdzk1 ‐KO ESCs and WT‐ESCs.

Article Snippet: The primary antibodies used were CDX2 (BioGenes, MU392A), OCT4 (Abcam, ab181557), GATA6 (CST, 5851), EOMES (Abcam, ab23345), TFAP2C (Santa Cruz, sc‐12762), and YAP1 (ABclonal, A1002).

Techniques: Expressing