Journal: Cell reports

Article Title: Paraspeckle protein NONO regulates active chromatin by allosterically stimulating NSD1

doi: 10.1016/j.celrep.2025.116247

Figure Lengend Snippet: (A) Schematic illustration of experimental flows for proteomics analysis of NSD1-PWWP2’s interactomes. (B) Unique proteins detected by LC-MS and plotted by peptide-spectrum match (PSM) scores against percentage of coverage using DIPG13 (top) and HEK293T (bottom) cells. (C) Illustration of annotated functional domains of NONO. (D) GST pulldown assay of HA-tagged NONO using NSD1-PWWP2 as the bait. Left, pulldown of HA-tagged N-NONO or C-NONO using GST alone or GST-NSD1-PWWP2 followed by western blot of GST and HA. Right, pulldown of HA-tagged N-NONO using GST-NSD1-PWWP2 or GST-NSD1-PWWP2–4A mutant followed by western blot of GST and HA.

Article Snippet: NONO rabbit polyclonal , ProteinTech , Cat# 11058-1-AP.

Techniques: Liquid Chromatography with Mass Spectroscopy, Functional Assay, GST Pulldown Assay, Western Blot, Mutagenesis

Journal: Cell reports

Article Title: Paraspeckle protein NONO regulates active chromatin by allosterically stimulating NSD1

doi: 10.1016/j.celrep.2025.116247

Figure Lengend Snippet: (A) Demonstration of recombinant protein expression and purification, including NSD1, NSD1 PWWP2–4A , N-NONO, and recombinant di-nucleosomes by Coomassie blue staining. (B) HMT assays of full-length NSD1 or NSD1 PWWP2–4A mutant in an incremental titration of 62.5, 125, and 250 nM. Top: quantifications of autoradiographic signals normalized to NSD1 alone (the second lane). Middle: representative autoradiographic images for stably incorporated [ 3 H]. Bottom: Coomassie blue staining of total nucleosomes. Data are presented as mean ± SEM. (C) HMT assays of 60 nM full-length NSD1 with an incremental titration of N-NONO at 0, 240, 530, 880, and 1760 nM. Top: quantifications of autoradiographic signals normalized to NSD1 alone (the second lane). Middle: representative autoradiographic images for stably incorporated [ 3 H]. Bottom: Coomassie blue staining of total nucleosomes. Data are presented as mean ± SEM. (D) HMT assays of 0.25 μM NSD1 PWWP2–4A mutant with an incremental titration of N-NONO at 0, 530, 880, and 1760 nM. Top: quantifications of autoradiographic signals normalized to NSD1 alone (the second lane). Middle: representative autoradiographic images for stably incorporated [ 3 H]. Bottom, Coomassie blue staining of total nucleosomes. Data are presented as mean ± SEM.

Article Snippet: NONO rabbit polyclonal , ProteinTech , Cat# 11058-1-AP.

Techniques: Recombinant, Expressing, Purification, Staining, Mutagenesis, Titration, Stable Transfection

Journal: Cell reports

Article Title: Paraspeckle protein NONO regulates active chromatin by allosterically stimulating NSD1

doi: 10.1016/j.celrep.2025.116247

Figure Lengend Snippet: (A) Overlay of meta-analysis profiling of H3K36me2 ChIP-seq signals at all genes within a window of −10 kb of TSS to +10 kb of TES in WT and NONO-KO E14-mESC. Representative track images are shown at the bottom. (B) Individual meta-analysis profiling and heatmaps of H3K36me2 ChIP-seq in WT and NONO-KO mESCs. Left: ChIP-seq signals from WT cells were presented at all genes within a −10 kb of TSS to +10 kb of TES window, and NONO-KO cells were aligned to WT cells. Right: ChIP-seq signals were ranked by max peak value and aligned to the centers. (C) qPCR quantification of NEAT1 RNA expression levels in WT and NEAT1 CRISPRi cells. Signals were normalized by GAPDH . n = 5 for each condition. p value was calculated by Student’s t test. Data are presented as mean ± SEM. (D) Immunofluorescence staining of NONO in WT and NEAT1 CRISPRi HEK293T cells. Images were captured under a 63× objective, and the puncta of nuclear paraspeckles were highlighted by red triangles. Scale bars, 50 μm. (E) Quantifications of (D). Nuclear paraspeckles are present in individual WT ( n = 24) and NEAT1 CRISPRi ( n = 40) HEK293T cells. The p value is calculated by chi-squared test. (F) Overlay of meta-analysis profiling of H3K36me2 ChIP-seq signals at all genes within a window of −10 kb of TSS to +10 kb of TES in WT and NEAT1 CRISPRi HEK293T cells. Representative track images are shown at the bottom. (G) Individual meta-analysis profiling and heatmaps of H3K36me2 ChIP-seq in WT and NEAT1 CRISPRi HEK293T cells. ChIP-seq signals from WT cells were presented at all genes within a −10 kb of TSS to +10 kb of TES window, and NONO-KO cells were aligned to WT cells. Right: ChIP-seq signals were ranked by max peak value and aligned to the centers.

Article Snippet: NONO rabbit polyclonal , ProteinTech , Cat# 11058-1-AP.

Techniques: ChIP-sequencing, RNA Expression, Immunofluorescence, Staining

Journal: Cell reports

Article Title: Paraspeckle protein NONO regulates active chromatin by allosterically stimulating NSD1

doi: 10.1016/j.celrep.2025.116247

Figure Lengend Snippet: (A) Meta-analysis profiling and heatmaps of NSD1 ChIP-seq signals at all genes within a window of −10 kb of TSS to +10 kb of TES in WT and NONO-KO HEK293T cells. NONO-KO is aligned to WT. (B) Meta-analysis profiling and heatmaps of NONO ChIP-seq signals at all genes within a window of −10 kb of TSS to +10 kb of TES in WT and NSD1-KO HEK293T cells. NSD1-KO is aligned to WT. (C) Meta-analysis profiling and heatmaps of NSD1 ChIP-seq signals at all genes within a window of −10 kb of TSS to +10 kb of TES in WT and NONO-KO mESC cells. NONO-KO is aligned to WT. (D) Meta-analysis profiling and heatmaps of NONO ChIP-seq signals at all genes within a window of −10 kb of TSS to +10 kb of TES in WT and NSD1-KO mESC cells. NSD1-KO is aligned to WT.

Article Snippet: NONO rabbit polyclonal , ProteinTech , Cat# 11058-1-AP.

Techniques: ChIP-sequencing

Journal: Cell reports

Article Title: Paraspeckle protein NONO regulates active chromatin by allosterically stimulating NSD1

doi: 10.1016/j.celrep.2025.116247

Figure Lengend Snippet: (A) Neural progenitor cell (NPC) differentiation of WT, NSD1-KO, and NONO-KO E14-mESCs. Top: representative images of embryoid bodies (EBs) undergoing NPC differentiation after 3 days of retinoic acid (RA) treatment. Bottom, quantifications of fully differentiated, partially differentiated, or non-differentiated EBs. Scale bars, 500 μm. (B) Heatmaps of differential gene expression analysis in WT, NSD1-KO, and NONO-KO cells treated with RA for 0, 3, or 6 days using RNA-seq. A total of 252 genes associated with neural development and 102 genes associated with stem cell differentiation were presented. (C) Heatmaps of significant changes of gene set enrichment analysis signatures, including stem cell differentiation and neural lineage gene sets in WT compared to NSD1-KO and NONO-KO E14-mESC cells undergoing RA-induced NPC differentiation. (D) Boxplots of log2 fold changes in gene expression using the experimental conditions shown in (B). The box and whisker represent 95%, the third quartile, the median, the first quartile, and 5% distribution of genes. Data are presented as mean ± SEM. p values were calculated by Wilcoxon test ** p < 0.01; *** p < 0.001; and **** p < 0.0001.

Article Snippet: NONO rabbit polyclonal , ProteinTech , Cat# 11058-1-AP.

Techniques: Gene Expression, RNA Sequencing, Cell Differentiation, Whisker Assay