Fig. 3.

Effect of TBX2 mutants on ADAM10 transcriptional activity. a Analysis of TBX2 repressor domain (RD) deletion mutant: schematic view of applied human TBX2 protein variants: DDK-tagged full-length TBX2 and respective protein sequence lacking the C-terminal part including the repressor domain (ΔRD). Expression of the TBX2 variants was verified by western blot method using both, TBX2 and DDK antibodies. Actin was detected for loading control. b TBX2 expression constructs were transfected together with ADAM10-promoter luciferase construct or promoter-less control plasmid in human SH-SY5Y cells. Luminescence values were normalized to protein content of respective cell lysate and values obtained for empty vector transfected cells set to 100%. c Effect of DNA-binding attenuated mutant of Tbx2 on ADAM10-promoter activity: schematic view of DDK-tagged murine full-length Tbx2 and the Tbx2 R122/123E DNA-binding impaired mutant (mTbx2_mut, mutation is indicated with a black triangle, both with a C-terminal DDK-Tag). Overexpression in murine N2a cells was analyzed by western blot and Actin served as loading control. A low molecular weight band occurring in Tbx2 overexpressing cells might represent a degradation product (band is marked with *). d N2a cells were transfected with respective Tbx2 expression plasmids together with murine Adam10-promoter reporter plasmid or promoter-less control vector. Values were normalized to protein content and values for control vector transfected cells were set to 100%. b, d Bars represent mean ± SD of three independent experiments conducted in triplicate (ns p > 0.05, *p < 0.05, One-way ANOVA, Bonferroni post-test)