Journal: Scientific reports

Article Title: Starvation-induced HBP metabolic reprogramming and STAM2 O-GlcNAcylation facilitate bladder cancer metastasis.

doi: 10.1038/s41598-025-92579-4

Figure Lengend Snippet: Fig. 2. STAM2 is upregulated in BCa cells and tissues. (A-B) STAM2 mRNA and protein levels were detected using qRT‒PCR and western blotting in tumor tissues and adjacent normal tissues. C Quantitative western blot analysis of the data in B. (D-E) STAM2 mRNA and protein levels were detected using qRT‒PCR and western blotting in an epithelial cell line (SV-HUC-1) and BCa cell lines (UM-UC-3, T24, and 5637). (F) Quantitative western blot analysis of E. (All PVDF bands were transferred and tested in the same western blot experiments, and the experiments performed in triplicate.)

Article Snippet: The antibodies used in the study were as follows: STAM2 (Proteintech, 13009-1-AP), HA (Proteintech,51064-2-AP), E-cadherin (Proteintech, 20874-1- AP), N-cadherin (Proteintech, 22018-1-AP), Vimentin (Proteintech, 10366-1-AP), JAK2 (CST, #3230), p-JAK2 (CST, #3771), stat3 (CST, #12640), p-stat3 (CST, #94994), and β-actin (Proteintech, 66009-1-Ig).

Techniques: Western Blot

Journal: Scientific reports

Article Title: Starvation-induced HBP metabolic reprogramming and STAM2 O-GlcNAcylation facilitate bladder cancer metastasis.

doi: 10.1038/s41598-025-92579-4

Figure Lengend Snippet: Fig. 4. STAM2 S375 O-GlcNAcylation enhances its stability by decreasing ubiquitination. Half-life and quantitative analysis of STAM2-HA (A-B) and STAM2 S375A-HA (C-D) in UM-UC-3 cells. E Co-IP assays detected the interaction between STAM2-HA and Ub-Flag in UM-UC-3 cells transfected with STAM2-HA. (F) Quantitative western blot analysis of ubiquitination. (G) Co-IP assays detected the interaction between STAM2-HA and Ub-Flag in UM-UC-3 cells transfected with STAM2 S375A-HA. (H) Quantitative western blot analysis of ubiquitination. (I) Immunofluorescence staining of STAM2-HA and OGT-Flag in UM-UC-3 cells treated with OSMI-1 or PUGNAc. (J) Quantification of the nuclear/cytoplasmic fluorescence ratios of STAM2. (All PVDF bands were transferred and tested in the same western blot experiments, and the experiments performed in triplicate.)

Article Snippet: The antibodies used in the study were as follows: STAM2 (Proteintech, 13009-1-AP), HA (Proteintech,51064-2-AP), E-cadherin (Proteintech, 20874-1- AP), N-cadherin (Proteintech, 22018-1-AP), Vimentin (Proteintech, 10366-1-AP), JAK2 (CST, #3230), p-JAK2 (CST, #3771), stat3 (CST, #12640), p-stat3 (CST, #94994), and β-actin (Proteintech, 66009-1-Ig).

Techniques: Ubiquitin Proteomics, Co-Immunoprecipitation Assay, Transfection, Western Blot, Immunofluorescence, Staining, Fluorescence

Journal: Scientific reports

Article Title: Starvation-induced HBP metabolic reprogramming and STAM2 O-GlcNAcylation facilitate bladder cancer metastasis.

doi: 10.1038/s41598-025-92579-4

Figure Lengend Snippet: Fig. 5. STAM2 S375 O-GlcNAcylation facilitates the JAK2–STAT3 signaling pathway. (A) STAM2 protein levels were measured using western blotting. (B) JAK2/p-JAK2 and STAT3/p-STAT3 protein levels were measured using western blotting in NC cells, STAM2-overexpressing cells and STAM2-knockout cells. (C) Quantitative western blot analysis of B. (D) JAK2/p-JAK2 and STAT3/p-STAT3 protein levels were measured using western blot in UM-UC-3 cells transfected with STAM2-HA or STAM2 S375-HA and treated with PUGNAc or not. (E) Quantitative western blot analysis of D. (F-G) Proliferation, migration, and invasion were measured using CCK-8 and Transwell assays in NC cells or cells transfected with STAM2-HA or STAM2 S375-HA after treatment with JSI-124. (H) EMT marker protein levels were measured using western blotting. (I) Quantitative western blot analysis of H. All PVDF bands were transferred and tested in the same western blot experiments. All the results represent three distinct repetitions. Statistical significance is denoted as ***p < 0.001, **p < 0.01, and *p < 0.05, indicating significant differences between two groups.

Article Snippet: The antibodies used in the study were as follows: STAM2 (Proteintech, 13009-1-AP), HA (Proteintech,51064-2-AP), E-cadherin (Proteintech, 20874-1- AP), N-cadherin (Proteintech, 22018-1-AP), Vimentin (Proteintech, 10366-1-AP), JAK2 (CST, #3230), p-JAK2 (CST, #3771), stat3 (CST, #12640), p-stat3 (CST, #94994), and β-actin (Proteintech, 66009-1-Ig).

Techniques: Western Blot, Knock-Out, Transfection, Migration, CCK-8 Assay, Marker

Journal: Scientific reports

Article Title: Starvation-induced HBP metabolic reprogramming and STAM2 O-GlcNAcylation facilitate bladder cancer metastasis.

doi: 10.1038/s41598-025-92579-4

Figure Lengend Snippet: Fig. 6. STAM2 S375 O-GlcNAcylation promotes tumor proliferation and metastasis in vivo. (A) Schematic representation of the establishment of UM-UC-3-derived xenografts. (B) Representative images of xenograft tumors 21 days after subcutaneous injection. C Tumor volumes were measured every 3 days. (D) Weights of xenograft tumors 21 days after subcutaneous injection. (E-H) Protein levels of EMT marker JAK2/p-JAK2 and STAT3/p-STAT3 were measured using western blotting. All PVDF bands were transferred and tested in the same western blot experiments, and the experiments performed in triplicate. (I-L) Bioluminescence detection and bioluminescence quantitative analysis of the lung (I-J) and popliteal lymph nodes (K-L). (M- N) Representative image of the popliteal lymphatic metastasis model, along with quantitative analysis of the volume of popliteal lymph nodes.

Article Snippet: The antibodies used in the study were as follows: STAM2 (Proteintech, 13009-1-AP), HA (Proteintech,51064-2-AP), E-cadherin (Proteintech, 20874-1- AP), N-cadherin (Proteintech, 22018-1-AP), Vimentin (Proteintech, 10366-1-AP), JAK2 (CST, #3230), p-JAK2 (CST, #3771), stat3 (CST, #12640), p-stat3 (CST, #94994), and β-actin (Proteintech, 66009-1-Ig).

Techniques: In Vivo, Derivative Assay, Injection, Marker, Western Blot