Figure 5.

STAT3 is involved in the transcriptional activation of S100A7 by IGF-1. (A) IGF-1 (10 nM, 24 h) induces the recruitment of STAT3 (Y705) to the STAT3 binding site located within the S100A7 promoter sequence, as ascertained by Chromatin Immunoprecipitation (ChIP)-RT-PCR assay performed in MCF-7 cells. In control samples, nonspecific IgG was used instead of the primary antibody. Data shown are the mean ± SEM of at least two independent experiments performed in duplicate. (B) Representative immunoblots of HA, pSTAT3 (Y705), and STAT3 in MCF-7 cells which were transiently transfected with pcDNA3 and STAT3-HA for 48 h, to generate a constitutively activated STAT3 system. (C) Evaluation of S100A7 mRNA expression by RT-PCR in MCF-7 cells transfected with pcDNA3 and STAT3-HA for 48 h, as indicated. 36B4 gene serves as normalizing control. (D) The transactivation of a S100A7 (pS100A7) promoter plasmid observed after treatment with IGF-1 (10 nM, 18 h) is enhanced in MCF-7 cells transfected for 48 h with STAT3-HA, compared with cells transfected with pcDNA3 control vector. Results are expressed as the % change of normalized RLU values relative to vehicle-treated cells. Data shown represent the mean ± SEM of at least two independent experiments performed in triplicate. (*), (#) p < 0.05; (**) p < 0.01; (***) p < 0.001. Side panel shows a schematic representation of human S100A7 promoter carrying the STAT3-responsive site.