Fig. 5: KY216 inhibits EMT by promoting VASH2 binding to α-tubulin.

a deTyr-tub protein blot analysis in NCI-H460 and 95-D cells (independant biological replicates n = 3). b Immunofluorescence analysis of deTyr-tub (green), cell nuclei were stained with DAPI (blue), and images were acquired by the confocal microscopy system (independant biological replicates n = 3). Scale bar: 20 μm. Data are shown as means ± SEM. Values shown are P values and were calculated using the one-way ANOVA with Tukey’s comparisons test. c Co-Immunoprecipitation (Co-IP) showed that the binding of VASH2 to α-tubulin in H460 cells was enhanced by KY216 (20 nM) (independant biological replicates n = 3). d The interactions between VASH2 and α-tubulin under KY216 treatment were detected using the PLA. Representative fluorescence images are shown (independant biological replicates n = 3). Scale bar: 20 μm. e Co-IP results showed that KY216 (20 nM) promoted the interaction of VASH2 with α-tubulin and enhanced the process of α-tubulin detyrosination (independant biological replicates n = 3). f Schematic depiction of VASH2. The Cys-His-Ser catalytic triad is highlighted in yellow circles. g Protein blotting was used to analyze changes in the expression of ZEB1 and deTyr-tub following transfection of VASH2 wild-type and mutant plasmids in 95-D cells (independant biological replicates n = 3). h Changes in the interaction of VASH2 (eGFP-tagged) with α-tubulin after transfection of VASH2 wild-type and mutant plasmids in 95-D cells were investigated by Co-IP analysis (independant biological replicates n = 3). i Protein blotting was utilized to detect alterations in the levels of ZEB1 and deTyr-tub in 95-D cells following transfection of VASH2-WT and VASH2-S210A mutant plasmids in the presence of 20 nM KY216 (independant biological replicates n = 3). j Co-IP assay was used to study changes in the interaction between VASH2 (eGFP-tagged) and α-tubulin after transfection of VASH2-WT and VASH2-S210A mutant plasmids in the presence of 20 nM KY216 in 95-D cells (independant biological replicates n = 3). Source data are provided as a Source Data file.