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Fig. 4 | Cell Division

Fig. 4

From: The T-box transcription factor TBX3 drives proliferation by direct repression of the p21WAF1 cyclin-dependent kinase inhibitor

Fig. 4

Phosphorylation of TBX3 at SP190 can affect its ability to repress p21. a Alignment showing conservation of the S190 residue (indicated as SP in red) of the TBX3 protein across a number of different species. b ATDC5 and SW1353 cells were co-transfected with a p21 promoter luciferase reporter construct together with pCMV empty, TBX3 WT, TBX3 S190A or TBX3 S190E expression constructs. The plasmid pRL-TK containing the Renilla luciferase reporter gene was also introduced to normalize transfection efficiency. Fold repression was obtained by comparing the results to that of the empty pCMV vector transfection. For luciferase analyses, the values indicate the mean of three independent experiments ± SEM (*p < 0.05 and ***p < 0.001). Lower panels western blots of transfected HA tagged-TBX3 proteins using an antibody to HA. c DNA-affinity immunoblot assay. Left panel lysates from SW1353 cells transfected with empty vector, TBX3WT, TBX3 S190A or TBX3 S190E constructs were incubated with a biotinylated probe matching the T-element at position -121 bp of the p21 promoter. A pull-down was performed using streptavidin magnetic beads and the lysates run on an 8 % SDS-PAGE gel and analysed by western blotting with an antibody to HA. Right panel western blotting to confirm the equal expression of the HA- tagged WT TBX3, TBX3 S190A and TBX3 S190E constructs using an antibody to HA

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