Figure 4

The transcription of TGF-βR2 is regulated by ZNF32. (a) An immunofluorescence assay shows stronger TGF-βR2 (green) expression in the Lv-ZNF32 group and weaker expression in the Sh-ZNF32 group. (b) Schematic representation of the ZNF32-binding sites in the TGF-βR2 promoter. The probe sequence is underlined, and the transcription start site is indicated by +1. (c) HEK293 cells were transiently transfected with TGF-βR2 promoter 5′-deletion mutant constructs and analyzed through a dual-luciferase reporter assay. (d) HEK293 cells were transiently transfected with the indicated constructs, treated as in (c) and then analyzed using a dual-luciferase reporter assay. (e) Nuclear extracts from A549 cells were incubated in biotin-labeled oligonucleotides corresponding to the TGF-βR2 promoter region −736/−751. The arrow shows the specific DNA-protein complex. (f) DNA fragments from A549 cells were immunoprecipitated with ZNF32-specific antibodies and analyzed via RT-PCR using the indicated primers. NS, non-significant difference. The data are presented as the means±S.D. Each experiment was performed at least in triplicate, and consistent results were obtained. The photograph shows a representative result from three independent experiments