Journal: Cell Death Discovery
Article Title: MYCN inhibits TrkC-mediated differentiation in neuroblastoma cells via disruption of the PKA signalling pathway
doi: 10.1038/s41420-026-03024-y
Figure Lengend Snippet: A MYCN expression of the SH-SY5Y, NBLS and NLF parental neuroblastoma cell lines by Western blotting. B TrkC and V5-tag expression in parental neuroblastoma cell lines and pLX302/NTRK3 transfected cells. C Western blot analyses of phospho-TrkC (Tyr 516 ), phospho-ERK (Thr 202 /Tyr 204 ), phospho-PLCγ (Tyr 783 ) following stimulation of SH-SY5Y/TrkC, NBLS/TrkC, NLF/TrkC cells with 100 ng/ml of NT-3 for 10 min. GAPDH acted as loading control for Western blot experiments. D Representative images of phenotypic observation by light microscopy 5 days post NT-3 (100 ng/ml) stimulation in SH-SY5Y/TrkC, NLF/TrkC and NBLS/TrkC neuroblastoma cells. Magnification: X20; scalebar: 200 µm. E Quantification of neuronal differentiation following NT-3 (100 ng/ml) treatment for 5 days. ( n = 3). F Western blot analysis of GAP43, a neuronal differentiation marker, following stimulation of SH-SY5Y/TrkC cells with 100 ng/ml of NT-3 for 5 days. Vinculin acted as loading control. ( n = 3). G Quantification of mean fluorescence as a measure of cell proliferation at 72 h post NT-3 (100 ng/ml) stimulation using CyQuant assay. Data shown as mean ± SEM; n = 3 ( p < 0.05 = *, p < 0.001 = **, p < 0.0001 = ***).
Article Snippet: Other antibodies used include TrkC (C44H5) (#3376), Phospho-ERK (Thr 202 /Tyr 204 ) (#4370), Phospho-Akt (Ser 473 ) (#4060), Akt (#9272), Phospho-PKA C (Thr 197 ) (#4781), PKA C-α (#4782), Vinculin (#4650), CREB (#9197), Phospho-CREB (Ser 133 ) (#9191), GAP43 (#8945), Tyrosine hydroxylase (#2792), GSK-3β (#12456), and GAPDH (14C10) (#2118), from Cell Signalling Technology.
Techniques: Expressing, Western Blot, Transfection, Control, Light Microscopy, Marker, Fluorescence, CyQUANT Assay
