human wnt3a protein  (R&D Systems)

Journal: The Journal of Biological Chemistry

Article Title: IFIT3-DVL interaction promotes malignant progression of lung squamous cell carcinoma and large-cell lung carcinoma via canonical WNT signaling

doi: 10.1016/j.jbc.2026.111368

Figure Lengend Snippet: Effect of IFIT3 on cell malignancy and canonical WNT signaling. A and B , Western blot analysis. GAPDH was an internal control. Quantitative analysis of band intensities is provided in . p-GSK-3β (Ser9), the serine nine phosphorylation of GSK-3β. C–H , quantitative real-time PCR analysis. β-actin was an internal control. Vec, vector, the control group of the IFIT3 expression plasmid. IF3, transfection of IFIT3 expression plasmid. si-IF3, siRNA-IFIT3. si-NC, siRNA-negative control, a scrambled siRNA selected as a negative control. si-IF3 (IF3), following IFIT3 knockdown with siRNA, its expression was restored by transfection with IFIT3 expression plasmid. I–K , dual-luciferase assay. IFIT3 expression plasmid or IFIT3 siRNA was transfected to LUSC/LCLC cells, with or without the treatment of recombinant WNT3A protein. (i) Negative Control: transfection of empty plasmid vector or siRNA-negative control. (ii) Positive Control: treatment of recombinant WNT3A protein. (iii) Test Group 1: transfection of IFIT3 expression plasmid or IFIT3 siRNA. (iv) Test Group 2: transfection of IFIT3 expression plasmid or IFIT3 siRNA, with the treatment of recombinant WNT3A protein. rhW3A, the cells were treated with recombinant human WNT3A protein (100 ng/ml) for 12 h. Error bars indicated mean ± SEM (n = 3). Data were analyzed by two-tailed unpaired Student’s t test for ( C , D , F and G ), and one-way ANOVA with Tukey’s post hoc test for ( E , H–K ). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001. LCLC, large-cell lung carcinoma; LUSC, lung squamous cell carcinoma.

Article Snippet: For analysis of WNT3A response, cells were treated with recombinant human WNT3A protein (100 ng/ml, R&D systems, Cat# 5036-WN-500/CF) for 12 h prior to dual-luciferase assay or Western blot analysis.

Techniques: Western Blot, Control, Phospho-proteomics, Real-time Polymerase Chain Reaction, Plasmid Preparation, Expressing, Transfection, Negative Control, Knockdown, Luciferase, Recombinant, Positive Control, Two Tailed Test

Journal: bioRxiv

Article Title: Nup358 Sustains Intestinal Epithelial Homeostasis by Preventing Dvl1 Condensate Formation to Restrain Wnt Signaling

doi: 10.64898/2026.03.25.714063

Figure Lengend Snippet: a. The expression levels of Wnt target genes Axin2, Ccnd1, and cMyc were analyzed in small intestinal epithelium from Nup358 fl/fl and Nup358 fl/fl CreER T 2 mice treated with tamoxifen by qPCR. Data are expressed as mean ± SD. *p ≤ 0.05 and **p ≤ 0.01. b. Immunofluorescence staining for TCF-4 in small intestinal epithelium from Nup358 fl/fl and Nup358 fl/fl CreER T 2 mice treated with tamoxifen. c. Fluorescent Wnt reporter activity in HEK293T cells transfected with either control or Nup358-specific siRNA and treated with either Wnt3a or vehicle were analyzed by confocal microscopy (top) and quantified by flow cytometry (bottom). Data are expressed as mean ± SD, ****p ≤ 0.0001. d. Fluorescent Wnt reporter activity in HEK293T cells transfected with control siRNA, β-catenin-specific siRNA, Nup358-specific siRNA, or a combination of Nup358-specific siRNA and β-catenin-specific siRNA and treated with either Wnt3a or vehicle were analyzed by confocal microscopy (top) and quantified (bottom). Data are expressed as mean ± SD. *p ≤ 0.05 and ****p ≤ 0.0001.

Article Snippet: Reporter activity was validated by treating the stable line with recombinant Wnt3a (100 ng/mL, Proteintech HZ-1296) and confirming EGFP induction via fluorescence microscopy.

Techniques: Expressing, Immunofluorescence, Staining, Activity Assay, Transfection, Control, Confocal Microscopy, Flow Cytometry

Journal: bioRxiv

Article Title: Nup358 Sustains Intestinal Epithelial Homeostasis by Preventing Dvl1 Condensate Formation to Restrain Wnt Signaling

doi: 10.64898/2026.03.25.714063

Figure Lengend Snippet: a. Levels of the 7TGP fluorescent Wnt reporter in HEK293T cells transfected with control siRNA, Dvl1-specific siRNA, Nup358-specific siRNA, or a combination of Nup358-specific siRNA and Dvl1-specific siRNA and treated with either Wnt3a or vehicle. b. Western blot analysis and relative quantification of protein levels of Dvl1 and Axin1 in HEK293T cells transfected with either control or Nup358-specific siRNA. α-Tubulin and GAPDH were used as loading controls. Data are expressed as mean ± SD. **p ≤ 0.01. c. Immunofluorescence staining for Dvl1 and Nup358 in HEK293T cells transfected with either control or Nup358-specific siRNA. d. Immunofluorescence staining for Dvl1 and Nup358 in HEK293T cells transfected with control or Nup358-specific siRNA and treated with vehicle or 1,6-hexanediol 5% for 2 minutes. e. Fluorescence recovery after photobleaching (FRAP) and fusion analysis of Dvl1 condensates in HEK293T cells transfected with Nup358-specific siRNA and EGFP-Dvl1.

Article Snippet: Reporter activity was validated by treating the stable line with recombinant Wnt3a (100 ng/mL, Proteintech HZ-1296) and confirming EGFP induction via fluorescence microscopy.

Techniques: Transfection, Control, Western Blot, Quantitative Proteomics, Immunofluorescence, Staining, Fluorescence

Journal: bioRxiv

Article Title: Nup358 Sustains Intestinal Epithelial Homeostasis by Preventing Dvl1 Condensate Formation to Restrain Wnt Signaling

doi: 10.64898/2026.03.25.714063

Figure Lengend Snippet: a. Levels of the 7TGP fluorescent Wnt reporter in HEK293T cells transfected with control siRNA, Dvl1-specific siRNA, Nup358-specific siRNA, or a combination of Nup358-specific siRNA and Dvl1-specific siRNA and treated with either Wnt3a or vehicle. b. Western blot analysis and relative quantification of protein levels of Dvl1 and Axin1 in HEK293T cells transfected with either control or Nup358-specific siRNA. α-Tubulin and GAPDH were used as loading controls. Data are expressed as mean ± SD. **p ≤ 0.01. c. Immunofluorescence staining for Dvl1 and Nup358 in HEK293T cells transfected with either control or Nup358-specific siRNA. d. Immunofluorescence staining for Dvl1 and Nup358 in HEK293T cells transfected with control or Nup358-specific siRNA and treated with vehicle or 1,6-hexanediol 5% for 2 minutes. e. Fluorescence recovery after photobleaching (FRAP) and fusion analysis of Dvl1 condensates in HEK293T cells transfected with Nup358-specific siRNA and EGFP-Dvl1.

Article Snippet: Reporter activity was validated by treating the stable line with recombinant Wnt3a (100 ng/mL, Proteintech HZ-1296) and confirming EGFP induction via fluorescence microscopy.

Techniques: Transfection, Control, Western Blot, Quantitative Proteomics, Immunofluorescence, Staining, Fluorescence

Journal: bioRxiv

Article Title: Nup358 Sustains Intestinal Epithelial Homeostasis by Preventing Dvl1 Condensate Formation to Restrain Wnt Signaling

doi: 10.64898/2026.03.25.714063

Figure Lengend Snippet: a. Schematic representing different functional domains of Nup358. b. Fluorescent Wnt reporter activity in HEK293T cells transfected with control siRNA, Nup358-specific siRNA, Ubc9-specific siRNA, or treated with the SUMOylation inhibitor 2-D08. c. Schematic representing different HA-tagged truncated forms of Nup358 that were transiently expressed in HEK293T cells. d. Fluorescent Wnt reporter activity in HEK293T cells transfected with either control or Nup358-specific siRNA alone or in combination with different HA-tagged truncated forms of Nup358 were analyzed by confocal microscopy (top) and quantified (bottom). Data are expressed as mean ± SD. ****p ≤ 0.0001 e. Western blot analysis of co-immunoprecipitation of HA-Nup358 and EGFP-Dvl1 transiently expressed in HEK293T cells. GAPDH was used as loading control. f. Immunofluorescence of HEK293T cells transfected with HA-Tagged Nup358 1–1133 and EGFP-Dvl1and stained with an anti-HA antibody. g. Immunofluorescence staining for endogenous Dvl1 and Nup358 in HEK293T cells transfected with either scramble control or Nup358-specific siRNA alone or in combination with HA-Tagged Nup358 1–1133 fragment were analyzed by confocal microscopy (top) and quantified (bottom). Data are expressed as mean ± SD. **p ≤ 0.01.

Article Snippet: Reporter activity was validated by treating the stable line with recombinant Wnt3a (100 ng/mL, Proteintech HZ-1296) and confirming EGFP induction via fluorescence microscopy.

Techniques: Functional Assay, Activity Assay, Transfection, Control, Confocal Microscopy, Western Blot, Immunoprecipitation, Immunofluorescence, Staining