FIGURE 3.

IL-1β-mediated increased αvβ8 expression in lung fibroblasts is dependent on ATF2 and SP3, but IL-1β did not increase their ability to bind to the ITGB8 core promoter oligonucleotides. A and B, lung fibroblasts were transfected with control and ATF2 (A) or SP3 (B) siRNA treated with or without IL-1β, and β8 expression was assessed using flow cytometry (n = 5). The results of pooled experiments are expressed as mean fluorescence intensity (MFI). siRNAs to SP3 and ATF2 efficiently knocked down expression of SP3 or ATF2, whereas treatment with the control siRNA had no effect (data not shown). *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001. C, EMSA of adult lung fibroblasts using a biotinylated probe (sequence indicated above) covering the CRE, CCAAT, and SP transcription factor binding sites in the core promoter with or without the indicated unlabeled competitors. WT is the unmutated competitor. CRE/CCAAT mt, SP, or triple mt indicates mutations in each or all of the indicated sites. CRE wt is an unrelated CRE consensus site competitor (24). Shown are data from two independent experiments with or without IL-1β treatment. In Experiment 1, the shifted SP3 (SP) and ATF2 (CRE) complexes are indicated by numbers and are identified by use of the non-labeled competitors (24). A fast migrating nonspecific band (ns) is indicated.