Journal: Nature Communications

Article Title: The chromatin, topological and regulatory properties of pluripotency-associated poised enhancers are conserved in vivo

doi: 10.1038/s41467-021-24641-4

Figure Lengend Snippet: A H3K27me3 HiChIP and ChIP-seq profiles generated in mESC are shown around the Lhx5 locus. Both the Lhx5 TSS and a PE previously shown to control this gene in AntNPC (PE Lhx5 (−109kb)) are marked with green vertical lines. Upper panel: heatmap of log2 interaction intensities based on H3K27me3 HiChIP data generated in mESC. Two medium panels: ChIP-seq and 1D HiChIP signals for H3K27me3 in mESC. Lower two panels: significant ( p < 0.01; FitHiChIP ) interactions were called using the H3K27me3 HiChIP PETs (paired-end tags) and loops in which one of the anchors overlaps (±10 kb) either the PE Lhx5 (−109kb) or the Lhx5 TSS are shown. b Significant ( p < 0.01; FitHiChIP) interactions were called using the mESC H3K27me3 HiChIP PETs (paired-end tags). Interaction anchors were overlapped with PEs and their interaction partners were hierarchically annotated with the categories shown in the pie chart (“Methods”). c mESC ChIP-seq profiles for H3K27me3, H3K27ac, and H3K4me3 are shown around the TSS ( n = 1083) interacting with PEs according to the H3K27me3 HiChIP loops identified in mESC ( p < 0.01; FitHiChIP). d Pathway (green) and Gene Ontology (red) analysis for the TSS/genes interacting with PEs according to the H3K27me3 HiChIP loops identified in mESC ( p < 0.01; FitHiChIP) was performed with ConsensusPathDB. e The interactions (called using mESC H3K27me3 HiChIP data; p < 0.01; FitHiChIP) established by PcG domains (“Methods”) and PcG-bound promoters as well as those involving PE-general promoter or PE-bivalent promoter pairs were classified as either intra-TAD (yellow) or inter-TAD (green) depending on whether the two anchors of each loop occur within the same or different TADs, respectively. f , g ) Significant (mESC H3K27me3 HiChIP data; p < 0.01; FitHiChIP) interactions between distal PEs (>10 kb from TSS) and bivalent promoters ( n = 526) were visualized as pileup plots using Hi-C data generated in vitro (2i ESC, serum+LIF ESC; f ) and in vivo (E3.5 Inner Cell Mass (ICM), E6.5 epiblast, E7.5 ectoderm; g ). Hi-C pairwise interactions are shown 250 kb up- and downstream of each PE and bivalent promoter pair. Hi-C interaction matrices were KR-balanced (balanced). The numbers in the upper left corners correspond to “loopiness” values (center intensity normalized to the intensity in the corners).

Article Snippet: Standard serum + LIF medium contained 500 mL Knockout DMEM (Gibco 10829-018), 95 mL of filtered ES FBS (Gibco 16141-061), 5.9 mL of antibiotics ( Hyclone SV30079.01 ), 5.9 mL Glutamax (Gibco 35050-038), 5.9 mL MEM NEAA (Gibco 11140-035), 4.7 mL titrated LIF (Miltenyi Biotec 130-095-777), and 1.3 mL Beta-mercaptoethanol 55 mM (Gibco 21985-023).

Techniques: HiChIP, ChIP-sequencing, Generated, Control, Hi-C, In Vitro, In Vivo

Journal: EMBO Molecular Medicine

Article Title: LIF, a mitogen for choroidal endothelial cells, protects the choriocapillaris: implications for prevention of geographic atrophy

doi: 10.15252/emmm.202114511

Figure Lengend Snippet: Q‐PCR (Taqman) analysis of LIF expression in BCE cells and bovine pericytes (bPericyte). Bovine LIF ELISA of BCE cells and bovine pericyte. Cells were cultured for 48 h before harvesting the medium. LIFR expression in BCE cells and bovine pericyte by qPCR (Taqman). Image of FACS selection of CD13 + /CD45 − and CD31 + /CD45 − cells. Immunostaining for CD31, CD13, and DAPI in isolated primary mouse cells before and after FACS selection. Mouse LIF expression in FACS‐selected cells. Human choroidal EC (HCEC) and mouse retinal EC (MREC) were treated with LIF 10 ng and LIF 100 ng for 30 min. (PBS was used as control). STAT3 phosphorylation was detected by Western blot. H&E staining of LIFR and CD31 in mouse eye sections. The magnified images of the dashed line boxes, a and b, are shown on the right panel. Box a shows LIFR staining in retina and box b showed LIFR staining in RPE/choroid layer. Whole‐mount staining of LIFR and CD31 in mouse retina. The fluorescent staining for CD31 (green) and LIFR (red) is shown. White arrows indicate the co‐staining of CD31 and LIFR on retinal ECs. Data information: Bars and error bars represent mean ± SEM. All experiments were carried out in three independent studies. Two‐way ANOVA was used as statistical test.

Article Snippet: Mouse and bovine LIF concentrations were measured using specific ELISA kits (R&D Systems, Cat# MLF00; MyBioSource, Inc, Cag# MBS766054).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Cell Culture, Selection, Immunostaining, Isolation, Western Blot, Staining