Fig 1: LINC00472 regulates H3K27 acetylation at the ITGB8 transcription start site.A Two significantly enriched histone modification peaks at the ITGB8 transcription start site in the ChIP-Seq database that is H3K4me3 (Top) and H3K27ac (Bottom). B Six pairs of primers were designed near the TSS site of ITGB8. C–F Chromatin immunocoprecipitation was used to detect the level of histone modification. Inhibition of LINC00472 expression significantly decreased the level of H3K27ac near ITGB8 TSS site (E), while the level of H3K4me3 not (C). Overexpression of LINC00472 distinctly increased the level of H3K27ac near ITGB8 TSS site (F), but the level of H3K4me3 not (D). G RNA FISH results showed that LINC00472 was distributed both in the nucleus and cytoplasm. H The total H3K27ac levels in 769-P and HK-2 cells were detected by Western Blot. I RNA immunocoprecipitation was used to verify the interaction between LINC00472 and EZH2 or P300. J Using the algorithm support provided by RPISeq and lncPro database to analyze the possibility of interaction between histone modification participants and LINC00472. K Prediction of interaction sites between LINC00472 and P300. L RNA FISH and immunofluorescence showed that LINC00472 and P300 had multiple overlaps in cells. In C–F and I, data were shown as mean ± SD of three independent experiments (*P < 0.05, **P < 0.01, ***P < 0.001, ns non-significant).
Fig 2: Schematic diagram of the proposed mechanism. EZH2 is enriched in the miR-125a-5p promoter and inhibits miR-125a-5p expression, thereby increasing the expression of the miR-125a-5p target SFMBT1. SFMBT1 further inhibits the expression of TGFβ1, and the extent of smad2 and smad3 phosphorylation (p-smad2/3), and consequently impedes keratinocyte apoptosis and promotes keratinocyte proliferation as well as the inflammatory reactions, eventually exacerbating psoriasis.EZH2, enhancer of zeste homolog 2; miR-125a-5p, microRNA-125a-5p; SFMBT1, Scm (Sex comb on midleg) with four MBT (malignant brain tumor) domains 1; TGFβ, transforming growth factor-β.
Fig 3: A schematic diagram of lncRNA TRERNA1 functions in DLBCL.ALKBH5-mediated upregulation of TRERNA1 with a low level of m6A modification can promote cell proliferation and cell cycle progression. The m6A modification of TRERNA1 was shown to epigenetically silence p21 transcription to accelerate cell cycle progression by interacting with EZH2.
Fig 4: PRMT5 associates with EZH2 in CRC cells. (A) Co-immunoprecipitation of endogenous EZH2 from HCT116 and SW480 cells overexpressing Flag-tagged PRMT5. IgG was used as the negative control. (B) Western blot analysis of EZH2 binding to purified GST and GST-tagged PRMT5 using EZH2 antibody (top). GST and GST-tagged PRMT5 from E. coli BL21 (DE3) strain were visualized by staining with Coomassie brilliant blue R-250 (bottom). The red asterisk and black asterisk were GST and GST-tagged PRMT5, respectively. (C) The subcellular location of endogenous PRMT5 and EZH2 proteins was analyzed in HCT116 and SW480 cells by immunofluorescence microscopy. Scale bar: 10 µm.
Fig 5: Methylation specific PCR of sFRP4 in SW480 cells. Inhibition of EZH2 in the SW480 colorectal cancer cells did not affect the hypermethylation status of sFRP4, and the promoter DNA methylation level was unchanged upon downregulation of EZH2. EZH2, enhancer of zeste homolog 2; M, methylation; U, unmethylation; MK, marker; NC, negative control; sFRP, secreted frizzle-related protein; si, small interfering.
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