shank3  (Cell Signaling Technology Inc)

Journal: bioRxiv

Article Title: Scaffold protein SHANK3 regulates endothelial cell motility and tissue mechanics

doi: 10.64898/2026.04.12.717721

Figure Lengend Snippet: A. UMAP of SHANK3 mRNA expression from single-cell transcriptome data (Tabula Sapiens ) shows high expression in endothelial cells. B. Histological staining of serial sections of human lung tissue. Insert shows SHANK3 expression in vein endothelial cells. PECAM-1 used as endothelial cell marker. Scale bar 100 µm. H&E, hematoxylin and eosin. C. Immunofluorescence images of HUVEC monolayers showing SHANK3 at linear cell junctions (white arrows) and protrusive structures that link to actin (orange arrows). Scale bar 20 µm. D. SHANK3 colocalises with cell junction components ZO-1, VE-cadherin (VE-cad) β-catenin and α-catenin. Yellow arrows indicate reticular adherens junctions. Scale bar 20 µm.

Article Snippet: TaqMan® Gene Expression Assays (Thermo Scientific) were used to detect Shank3 (Hs00873185_m1) and GAPDH (Hs02786624_g1).

Techniques: Expressing, Single Cell, Staining, Marker, Immunofluorescence

Journal: bioRxiv

Article Title: Scaffold protein SHANK3 regulates endothelial cell motility and tissue mechanics

doi: 10.64898/2026.04.12.717721

Figure Lengend Snippet: A. Immunofluorescence images showing subcellular localisation of myc-tagged BirA, BirA-SHANK3 and SHANK3-BirA in stably-expressing U2OS cells. Scale bar 20 µm. B. Interaction network of SHANK3 proximity interactors (BFDR ≤ 0.05). Nodes correspond to proximity interactors and edges indicate protein-protein interactions from STRING-db . BirA-SHANK3 interactors shown in green, SHANK3-BirA shown in blue, and shared interactors shown in turquoise. Square nodes indicate cell junction components (UniProt keyword ‘cell junction’; KW-0965) and thick outlines indicate previously identified SHANK3 interactors (BioGRID). C. GO analysis of SHANK3 proximity interactors (BFDR ≤ 0.05). The top 10 terms in each category are shown. D. SHANK3 proximity interactors with mRNA expression significantly up or down regulated in HUVECs in the presence or absence of laminar flow .

Article Snippet: TaqMan® Gene Expression Assays (Thermo Scientific) were used to detect Shank3 (Hs00873185_m1) and GAPDH (Hs02786624_g1).

Techniques: Immunofluorescence, Stable Transfection, Expressing, Protein-Protein interactions

Journal: bioRxiv

Article Title: Scaffold protein SHANK3 regulates endothelial cell motility and tissue mechanics

doi: 10.64898/2026.04.12.717721

Figure Lengend Snippet: A. Representative western blot and quantification of SHANK3 depletion in HUVECs using two siRNAs targeting SHANK3. GAPDH used as a loading control. Paired t-test. n = 3 biological replicates. B. Immunofluorescence images of HUVEC monolayers following siRNA depletion of SHANK3, showing SHANK3 depletion and altered cell morphology. Scale bar 50 µm. C. Quantification of cell shape from B using PECAM-1 to identify cell boundaries. Means per field of view shown. n = 3 biological replicates, 6 fields of view per condition each replicate. Unpaired t-test. D. Detection of gaps in the HUVEC monolayer using anti-fibronectin antibodies in non-permeabilised cells. Scale bar 50 µm. E. Quantification of fibronectin patches in D. n = 3 biological replicates, 10-13 fields of view per condition each replicate. Unpaired t-test. Control, AllStars negative control; ns, non-significant; ****, p value < 0.001; ***, p value < 0.005; **, p value < 0.01; *, p value < 0.05.

Article Snippet: TaqMan® Gene Expression Assays (Thermo Scientific) were used to detect Shank3 (Hs00873185_m1) and GAPDH (Hs02786624_g1).

Techniques: Western Blot, Control, Immunofluorescence, Negative Control

Journal: bioRxiv

Article Title: Scaffold protein SHANK3 regulates endothelial cell motility and tissue mechanics

doi: 10.64898/2026.04.12.717721

Figure Lengend Snippet: Retinal blood vessels (A, vascular front; B, artery; C, vein) from postnatal day 6 (P6) mice stained for Shank3 and VE-cadherin. White arrows indicate colocalization of Shank3 with VE-cadherin at endothelial cell–cell junctions in veins, capillaries, and arteries. Scale bars: 50 µm (A), 20 µm (B, C). D. Shank3 deletion was induced using 4-hydroxytamoxifen (4-OHT) in Shank3 flox/flox ( Shank3 iECKO ) and Shank3 flox/wt in tdTomato;Cdh5-CreER T2 background ( Shank3 iECKO/+ ) during postnatal days (P) 1–3, followed by analysis of retinal vasculature at P6 (n = 6–7 mice per genotype). Image generated in Biorender.com. E, F. Representative images of Isolectin B4 staining and tdTomato expression in retinal vasculature from Shank3 iECKO and Shank3 iECKO/+ pups. a, artery; v, vein. Scale bars: 200 µm (E), 500 µm (F). G. Quantification of sprouts per mm at P6 in the leading front from Shank3 iECKO and Shank3 iECKO/+ . H-K. Quantification of the retinal vasculature using the SproutAngio tool . Shown are total number of branches (H), radial expansion of the vasculature (I), skeleton length (J), and vessel density (K) at r = 0.5R and r = 0.7R, representing proximal and distal zones of the retinal vasculature with respect to optic nerve head (see Supplementary Fig. 8E). Results were normalised to the Shank3 iECKO/+ within litter. L. Representative images of sprouts and filopodia expressing tdTomato from Shank3 iECKO and Shank3 iECKO/+ control pups given 4-OHT at P2–3, followed by analysis of retinal vasculature at P6. Scale bar 20 µm. M. Analysis of total sprout length (n = 4–6 mice, described in L). N. Sprout length distribution (% of total) showing decreased longer sprouts in Shank3 iECKO compared to Shank3 iECKO/+ control. n = 4–6 mice (described in L). Statistical tests: two-tailed unpaired t-test (G-K, M), two-way ANOVA with Tukey’s multiple comparison (N). Data are presented as mean ± SEM. P values < 0.05 are shown. ***, p value < 0.005; *, p value < 0.05.

Article Snippet: TaqMan® Gene Expression Assays (Thermo Scientific) were used to detect Shank3 (Hs00873185_m1) and GAPDH (Hs02786624_g1).

Techniques: Staining, Generated, Expressing, Control, Two Tailed Test, Comparison

Journal: Journal of Neurochemistry

Article Title: Unveiling the Molecular Mechanism of Intestinal Metabolite para ‐Cresol in Modulating Neuroinflammation and Synaptic Dysfunction: Implications for Autism Spectrum Disorder

doi: 10.1111/jnc.70457

Figure Lengend Snippet: Shank3 knockdown in rat hippocampal neurons. Hippocampal neurons were transduced with GFP‐tagged Shank3 lentiviral shRNA particles on DIV1, then treated with vehicle or 100 μM p ‐Cresol from DIV8 to DIV14. Cells were immunostained with anti‐GFP, ‐Shank3, ‐VGLUT, and ‐VGAT Abs at the end of treatment. (A–F) Representative images of GFP + neurons (green) stained with anti‐Shank3 (magenta) in Scr (full expression of Shank3) and KD conditions (reduced expression of Shank3). Cell nuclei were counterstained with DAPI (blue). Scale bar = 20 μm. (G) Transfection efficiency of Shank3 KD lentiviral particles expressed as Shank3 puncta/100 μm. (H–W) Representative images of GFP + (green) Shank3 Scr and KD neurons stained with VGLUT (red) and VGAT (cyan) in ctrl and 100 μM p ‐Cresol conditions. Cell nuclei were counterstained with DAPI (blue). Scale bar = 20 μm. (X) VGLUT + and VGAT + puncta / 100 μm in Scr and Shank3 KD conditions, treated with vehicle or 100 μM p ‐Cresol. (Y) Effect of Shank3 KD combined with p ‐Cresol treatment on neuronal dendrite length (μm). All data were expressed as mean ± SEM values and analyzed by two‐way ANOVA followed by Tukey's post hoc comparisons: * p < 0.05; ** p < 0.01; *** p < 0.001 vs. Scr ctrl; £ p < 0.05; ££ p < 0.01; £££ p < 0.001 vs. Shank3 KD Ctrl; +++ p < 0.001 vs. Scr p ‐Cres 100 μM; #0.10 > p > 0.05.

Article Snippet: Cells were transduced with green fluorescent protein (GFP)‐tagged SH3 and multiple ankyrin repeat domains 3 (Shank3) rat shRNA lentiviral particles (ORIGENE, #TL710469V) on day in vitro (DIV) 1, with a multiplicity of infection of 5 for 20 h (hr).

Techniques: Knockdown, Transduction, shRNA, Staining, Expressing, Transfection