Journal: iScience

Article Title: A role for nucleolin in functional improvement in stroke

doi: 10.1016/j.isci.2026.116134

Figure Lengend Snippet: Ischemic cortical stroke increases nucleolin expression in excitatory neurons (A) Representative coronal section stained with DAPI with stroke (red dashed line) and area of interest (yellow box). (B) Representative in situ hybridization images of DAPI (nuclear counterstain), Slc17a7 (excitatory cortical neurons), Gad1 (inhibitory neurons), and nucleolin at time points across acute to chronic stroke. The dashed box indicates the region shown at higher resolution and magnification in accompanying panels. Scale bars, 20 μm, n = 5 animals per time point. (C) Quantification of nucleolin mRNA expression in excitatory cortical neurons post-stroke. Stroke induces both early and chronic expression of nucleolin in Slc17a7 + cells. 1-way ANOVA with post-hoc Tukey correction. Data are represented as mean ± SEM with data points representing individual animals. ∗ p = 0.039, ∗∗∗ p = 0.003 (3D post stroke), p = 0.004 (7D post stroke). (D) Quantification of nucleolin mRNA expression in inhibitory cortical neurons post-stroke. No significant change in nucleolin expression was detected in Gad1 + cells. Data are represented as mean ± SEM with data points representing individual animals. (E) Representative images of nucleolin protein expression in sham stroke and 7 days post-stroke mice. Scale bars, 10 μm, n = 5 animals per group. (F) Quantification of nucleolin protein expression post-stroke. Stroke significantly increases nucleolin expression 7 days post-stroke ( p = 0.0005 via t test). Data are represented as mean ± SEM with data points representing individual animals.

Article Snippet: Nucleolin heterozygous knockout mice , Doron-Mandel et al. , Jackson Laboratories , JAX 037120.

Techniques: Expressing, Staining, In Situ Hybridization

Journal: iScience

Article Title: A role for nucleolin in functional improvement in stroke

doi: 10.1016/j.isci.2026.116134

Figure Lengend Snippet: Heterozygous deletion of the nucleolin GAR domain is detrimental to post-stroke axonal sprouting (A) Representative in situ hybridization images show reduced nucleolin expression in GAR +/- mice. The dashed box indicates the region shown at higher resolution and magnification in accompanying panels. Scale bars, 20 μm, n = 5 animals per group. (B) Quantification of nucleolin mRNA in GAR wild type (GAR +/+ ) and heterozygous deletion (GAR +/- ) mice. GAR +/- mice show a significant reduction in nucleolin mRNA puncta in excitatory cortical neurons ( p = 0.004 via t test). Data are represented as mean ± SEM with data points representing individual animals. (C) Schematic shows the general surgery procedure. Generated via BioRender. (D) Quantitative cortical mapping of peri-infarct axonal sprouting in GAR heterozygous deletion mice compared to wild type littermates. Injection site noted via white circle. Stroke site indicated via black symbol. Heterozygous deletion of the nucleolin GAR domain significantly decreases peri-infarct axonal sprouting when compared to wild-type littermate controls ( p = 0.006 via Hotelling’s T2 test). (E) Axonal puncta distribution within concentric rings centered at the injection origin (0,0). Heterozygous deletion of the nucleolin GAR domain significantly reduces axon detection at distances greater than 3 mm from the injection site (∗ p = 0.0329, ∗∗ p = 0.00286, and ∗∗∗∗ p < 0.0001, general linear mixed model with post-hoc Tukey’s correction). Data are represented as mean ± SEM. (F) Representative cortical flat-map images of wild-type and heterozygous GAR deletion mice 1 month following cortical stroke. White circle indicates the tracer injection site, and the red dashed circle indicates the outer bounds of infarct. (G) Heterozygous deletion of the nucleolin GAR domain results in an approximately 2-fold increase in infarct size one month post stroke ( p = 0.0078 via t test). n = 5 animals per group for all measurements. Data are represented as mean ± SEM with data points representing individual animals.

Article Snippet: Nucleolin heterozygous knockout mice , Doron-Mandel et al. , Jackson Laboratories , JAX 037120.

Techniques: In Situ Hybridization, Expressing, Generated, Injection

Journal: iScience

Article Title: A role for nucleolin in functional improvement in stroke

doi: 10.1016/j.isci.2026.116134

Figure Lengend Snippet: GAR-induced somatic sequestration of nucleolin enhances post-stroke axonal sprouting (A) Quantitative cortical mapping of post-stroke axonal sprouting in flattened mouse cortex ipsilateral to the stroke. Exogenous GAR-induced somatic sequestration of nucleolin, injection site noted by the white circle, results in a significant increase in axonal sprouting to the adjacent somatosensory cortex compared to Dendra2 control virus ( p = 0.02727 via Hotelling’s T2 test). Stroke indicated via a black symbol. (B) Axonal puncta distribution within concentric rings centered at the injection site (0,0). Somatic sequestration of nucleolin significantly increases axon detection locally and distant from the injection site (∗∗ p = 0.00234, ∗∗∗ p = 0.000486, and ∗∗∗∗ p < 0.0001, general linear mixed model with post-hoc Tukey’s correction). Data are represented as mean ± SEM. (C) Representative flatmap images of mice injected with control and GARWT viruses. The white circle indicates the virus injection site, and the red dashed circle indicates the border of the infarct. Yellow square indicates zoomed in image, showing colocalization of Flag and NeuN in virally transduced cells. Scale bars, 20 μm, n = 5–7 animals per group. (D) Quantification of infarct size. Somatic sequestration of nucleolin does not impact infarct size ( p = 0.6989 via t test). n = 7 animals per group for all measurements. Data are represented as mean ± SEM with data points representing individual animals.

Article Snippet: Nucleolin heterozygous knockout mice , Doron-Mandel et al. , Jackson Laboratories , JAX 037120.

Techniques: Injection, Control, Virus

Journal: iScience

Article Title: A role for nucleolin in functional improvement in stroke

doi: 10.1016/j.isci.2026.116134

Figure Lengend Snippet: Nucleolin perturbation enhances post-stroke functional recovery (A) Diagram of grid-walking and pasta matrix tasks, and experimental timeline for task training and assessment post-stroke. (B) Somatic sequestration of nucleolin accelerates recovery in the pasta matrix fine motor control and grid-walking gross motor control tasks. Mice were evaluated for fine motor dexterity as defined by the percentage of baseline pieces of pasta broken. One-week post-stroke, both GARWT + stroke and Control + stroke mice showed a significant deficit in the number of pieces of pasta broken compared to Control alone (∗Control vs. Stroke + Control p = 0.0082, # Control vs. Stroke + GARWT p = 0.0003 via 2-way ANOVA). Three weeks post-stroke, Control + stroke mice continued to show a deficit, whereas GARWT + stroke mice had recovered to a degree with no significant difference from both control groups (∗ Control vs. Control + stroke p = 0.0021, Control vs. GARWT + stroke p = 0.4568, $ Control + stroke vs. GARWT + stroke p = 0.0280 via 2-way ANOVA). By five weeks post-stroke, both groups showed no significant difference from the control groups. N = 8–10 per group. Mice were evaluated for gross motor control as defined by the fold change of baseline percentage right foot faults. One-week post-stroke, Control + stroke and GARWT + stroke mice showed a significant increase in the relative number of foot faults involving the affected limb (∗Control vs. Control + stroke p = 0.0180, # Control vs. GARWT + stroke p = 0.0202 via 2-way ANOVA). At the endpoint of the experiment, 8 weeks post-stroke, Control + stroke mice showed a continued fold change increase in percent foot faults, whereas GARWT + stroke mice were not significantly different from Control alone (∗Control vs. Control + stroke p = 0.0323, Control vs. GARWT + stroke p = 0.3322). n = 12–15 per group. Data are represented as mean ± SEM. (C) Representative DAPI-stained cortical sections of stroke mice treated with control or GARWT virus. Yellow box outlines the peri-infarct region imaged for tissue-level outcomes, red dashed circle indicates infarct bounds. Scale bars, 20 μm. (D) Somatic sequestration of nucleolin has no effect on infarct size or tissue loss. n = 8 per group. p < 0.0001, control vs. control + stroke and control vs. GARWT + stroke via 2-way ANOVA with Tukey’s post-hoc correction. Data are represented as mean ± SEM with data points representing individual animals. (E) Somatic sequestration of nucleolin does not have an effect on Iba1 intensity following cortical stroke. n = 8 per group. Data are represented as mean ± SEM with data points representing individual animals.

Article Snippet: Nucleolin heterozygous knockout mice , Doron-Mandel et al. , Jackson Laboratories , JAX 037120.

Techniques: Functional Assay, Control, Staining, Virus

Journal: Cell Death & Disease

Article Title: The EHMT2-MBLAC2 axis suppresses ribosomal DNA transcription in response to nucleolar DNA damage

doi: 10.1038/s41419-026-08616-1

Figure Lengend Snippet: A–C HeLa I- Ppo I cells were pre-treated with ATM inhibitor (ATMi; KU-55933) or EHMT2 inhibitor (EHMT2i; UNC0638) for 1 h prior to I- Ppo I induction for 4 h. After fixation, cells were subjected to immunofluorescence with anti-C23 ( A ), anti-Ki-67 ( B ), anti-UBF ( C ) or anti-γH2AX antibodies, respectively. The dashed circles outline margins of the nuclei. Percentages of cells with the indicated protein-containing nucleolar caps were quantified from three experiments. D – F EHMT2-KO HeLa I- Ppo I cells were processed for immunofluorescence as described in ( A – C ). Cells were labeled with anti-C23 ( D ), anti-Ki-67 ( E ), anti-UBF ( F ) or anti-γH2AX antibodies, respectively. Nuclei were counterstained with DAPI. Percentages of cells with the nucleolar caps were quantified from three experiments. Bars represent mean ± SEM; ns not significant; * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: Antibodies used for immunofluorescence (IF) staining: anti-γH2AX (EMD Millipore, #JBW301); Anti-53BP1 (Novusbio, #NB100-304); Fibrillarin(CST, 2639T); anti-Ki-67 (Abcam, #ab15580); anti-C23 (Nucleolin; CST, #14574); anti-UBF (F-9) (Santa Cruz, #sc-13125); anti-BRCA1 (Santa Cruz, #sc-6954); Anti-Rad51 (Santa Cruz, # sc-398587); anti-Myc (Abclonal, #AE070); anti-GFP (Proteintech, #66002-1); Alexa Fluor 594 AffiniPure Goat Anti-Mouse IgG (H + L) (Jackson ImmunoResearch, #115-585-166); Alexa Fluor 488 AffiniPure Goat Anti-Rabbit IgG (H + L) (Jackson ImmunoResearch, #111-545-003); Alexa Fluor 488 AffiniPure Goat Anti-Mouse IgG (H + L) (Jackson ImmunoResearch, #115-545-003); Alexa FluorTM Plus 405 Anti-Rabbit (Invitrogen, # A48254 ); Alexa FluorTM Plus 405 Anti-Mouse (Invitrogen, #A48255).

Techniques: Immunofluorescence, Labeling

Journal: Cell Death & Disease

Article Title: The EHMT2-MBLAC2 axis suppresses ribosomal DNA transcription in response to nucleolar DNA damage

doi: 10.1038/s41419-026-08616-1

Figure Lengend Snippet: A The cellular localization of MBLAC2 in HeLa I- Ppo I cells following rDNA DSBs induction. HeLa I- Ppo I cells were transfected with EGFP-MBLAC2 and were treated with Shield-1 and 4-OHT for 4 h. Fixed cells were labeled with anti-53BP1, anti-γH2AX or anti-C23 antibodies. Nuclei were counterstained with DAPI. Enlarged images show the details of the indicated proteins. Quantification of relative signal intensities of EGFP-EHMT2, 53BP1, γH2AX and C23 was performed by ImageJ. The white lines in the enlargements of the representative images indicate the lines for analysis. The edge of the indicated nucleolar caps was labeled with dotted circle. B Analysis of nucleolar transcription activity by EU incorporation assay in MBLAC2-inactivated HeLa I- Ppo I cells following rDNA DSBs induction. Cells transduced with control (shCTR) or two independent MBLAC2-targeted shRNAs were induced for rDNA DSBs for 4 h. EU nucleolar intensity was subsequently determined by EU incorporation assay. At least 200 cells exhibiting well-circumscribed nucleoli were quantitatively assessed across two independent experiments. Quantification of relative EU nucleolar intensity is shown in Tukey boxplots. C Nucleolar EU intensities were analyzed in HeLa I- Ppo I cells treated with the EHMT2 gRNA or MBLAC2 siRNA after I- Ppo I induction. Relative nucleolar EU intensity was quantified from at least two independent experiments. Immunoblot of MBLAC2 and EHMT2 in the HeLa I- Ppo I cells induced with the indicated gRNA or siRNA. D Colony survival of HeLa I- Ppo I cells transfected with CTR siRNA and siRNA targeting MBLAC2 or EHMT2 following I- Ppo I induction, respectively. Cells were induced for rDNA DSBs for 5 min. After washing with PBS twice, cells were allowed to grow for two weeks before harvest and Coomassie blue staining. The relative outgrowth of the colonies between groups were quantified and plotted. The protein expression of MBLAC2 and EHMT2 in HeLa I- Ppo I cells transfected with the indicated siRNAs were examined by immunoblot. E Immunoblot of MBLAC2 and EHMT2 in the HeLa I- Ppo I cells transduced with control gRNA (CTR gRNA) and two EHMT2 gRNAs (EHMT2 KO1 and EHMT2 KO2). F shRNA-mediated MBLAC2 knockdown efficiency was measured by Western blotting. G The EHMT2-MBLAC2 interaction was confirmed by Co-immunoprecipitation (Co-IP). Flag-DYRK1B was used as the positive control. H HeLa I- Ppo I cells transduced with control gRNA (CTR gRNA) and two EHMT2 gRNAs (EHMT2 KO1 and EHMT2 KO2) were treated with cycloheximide for 0, 5, and 10 h. The protein expression of MBLAC2 and EHMT2 in HeLa I- Ppo I cells was examined by immunoblot. I The relative MBLAC2 protein level in HeLa I-PpoI cells treated with cycloheximide as depicted in ( A ) was measured. Data were derived from three independent experiments. J HeLa I-PpoI cells were transduced with control gRNA (CTR gRNA) and two EHMT2 gRNAs (EHMT2 KO1 and EHMT2 KO2). Fold change of MBLAC2 mRNA in I- Ppo I cells treated with cycloheximide was determined by RT-qPCR. Quantification of MBLAC2 mRNA fold change was from three independent experiments. K Proposed working model of EHMT2-MBLAC2 axis in promoting rDNA DSB-induced transcriptional suppression. Bars represent mean ± SEM; ns not significant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001.

Article Snippet: Antibodies used for immunofluorescence (IF) staining: anti-γH2AX (EMD Millipore, #JBW301); Anti-53BP1 (Novusbio, #NB100-304); Fibrillarin(CST, 2639T); anti-Ki-67 (Abcam, #ab15580); anti-C23 (Nucleolin; CST, #14574); anti-UBF (F-9) (Santa Cruz, #sc-13125); anti-BRCA1 (Santa Cruz, #sc-6954); Anti-Rad51 (Santa Cruz, # sc-398587); anti-Myc (Abclonal, #AE070); anti-GFP (Proteintech, #66002-1); Alexa Fluor 594 AffiniPure Goat Anti-Mouse IgG (H + L) (Jackson ImmunoResearch, #115-585-166); Alexa Fluor 488 AffiniPure Goat Anti-Rabbit IgG (H + L) (Jackson ImmunoResearch, #111-545-003); Alexa Fluor 488 AffiniPure Goat Anti-Mouse IgG (H + L) (Jackson ImmunoResearch, #115-545-003); Alexa FluorTM Plus 405 Anti-Rabbit (Invitrogen, # A48254 ); Alexa FluorTM Plus 405 Anti-Mouse (Invitrogen, #A48255).

Techniques: Transfection, Labeling, Activity Assay, Transduction, Control, Western Blot, Staining, Expressing, shRNA, Knockdown, Immunoprecipitation, Co-Immunoprecipitation Assay, Positive Control, Derivative Assay, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Nucleoli as drivers of nuclear remodelling in cardiomyocytes during Heart Failure

doi: 10.64898/2026.02.24.707499

Figure Lengend Snippet: Ai , Left, representative immunocytochemistry micrographs of detyrMT (green), actin (red) and nuclei (HOECHST, blue). White arrow indicates detyrMT inside the NI. Right, scatter plot of NI quantification in control, PTL and CytD-treated cardiomyocytes (N=3-5 rats, 44-106 nuclei). Aii , Line-scan schematic of cytoplasmic and nucleoplasmic CaTs in a cardiomyocyte. Aiii , Representative recordings of distinct subcellular regions, as indicated in (ii), baseline quantification in control, PTL and CytD at 1 and 2.5Hz (bottom) (N=3-4 rats, 32-36 cells). Bi , Left, representative immunocytochemistry micrographs of nucleolin (yellow) and nuclei (HOECHST, blue). White arrow indicates NI-nucleoli localization. Right, scatter plots of area, circularity (N=5 rats,162-209 nucleoli) and NI number (N=3 rats, 60-57 nuclei) in control and BMH21-treated cells. Ci , Left, representative immunohistochemistry micrographs α-actinin (red) and nuclei (HOECHST, blue). Right, scatter plot of NI quantification in donor and DCM tissue (N=8 patients, 99-120 cells). Di , Animal model schematic. Representative immunocytochemistry micrographs of nuclei (HOECHST, grey) in Dii and detyrMT (grey) in Diii . Scatter plots of NI quantification ( Dii ) in control, 8-weeks and 16-weeks MI cardiomyocytes (N=3-4 rats, 40-44 cells) and detyrMT area quantification in control and 8-weeks MI cardiomyocytes (N=3 rats, 35-42 cells). Div , Left, mechano-Scanning Ion Conductance Microscopy representative topography and Young’s modulus maps. Right, Young’s modulus quantification in control and 8-weeks MI (N=3 rats, 33-36 cells). Representative immunocytochemistry micrographs of nucleolin (magenta) and nuclei (HOECHST, blue) in Ei and nucleophosmin (magenta), γH2AX-DNA damage (green) and nuclei (HOECHST, blue) in Eii . Scatter plots of circularity (N=3-4 rats, 96-137 nucleoli) and chromocenter Manders Coefficient (M1,M2) (N=3 rats, 43-49 cells) in control and MI cardiomyocytes ( Ei ) and γH2AX foci area ( Eii ) (N=3 rats, 48-63 cells). Eiii , Left, representative heatmap. Right, Brillouin shift absolute difference in control and 8-weeks cardiomyocytes (N=3 rats, 36-37 cells). Statistics: nested t-test ( Bi , Diii-iv ), unpaired t-test Kolmogorov-Smirnov ( Bi , Ci , Ei-iii ), non-parametric Kruskal Wallis with Dunn’s multiple comparison ( Ai , Dii , Ei ).Normality tested with Shapiro-Wilk.Data represent mean±SEM. Scale bar 5µm.

Article Snippet: Primary antibodies used: anti-mouse α-actinin (Sigma, MA1-22863), anti-rabbit detyrosinated tubulin (Invitrogen, MA5-44591), anti-rabbit nucleolin (Cell Signaling, #14574), anti-mouse nucleophosmin (Abcam, ab10530), γH2AX (Cell Signaling, #5438).

Techniques: Immunocytochemistry, Control, Immunohistochemistry, Animal Model, Microscopy, Comparison