IL-13 is a central mediator of allergic inflammation. The single nucleotide polymorphism IL13-1112C>T (rs1800925) is associated with allergic phenotypes in ethnically distinct populations, but the underlying mechanism(s) remain unknown. Using in vivo, in vitro, and in silico analysis, we show that the IL13-1112T allele enhanced IL13 promoter activity in primary human and murine CD4+ Th2 lymphocytes. Increased expression of IL13-1112T in Th2 cells was associated with the creation of a Yin-Yang 1 binding site that overlapped a STAT motif involved in negative regulation of IL13 expression and attenuated STAT6-mediated transcriptional repression. Because IL-13 secretion was increased in IL13-1112TT homozygotes, we propose that increased expression of IL13-1112T in vivo may underlie its association with susceptibility to allergic inflammation. Interestingly, IL13-1112T had opposite transcriptional effects in nonpolarized CD4+ T cells, paralleled by distinct patterns of DNA-protein interactions at the IL13 promoter. Our findings suggest the nuclear milieu dictates the functional outcome of genetic variation.

The Th2 cytokine IL-13 is a major effector molecule in human allergic inflammation. Notably, IL-13 expression at birth correlates with subsequent susceptibility to atopic disease. In order to characterize the chromatin-based mechanisms that regulate IL-13 expression in human neonatal CD4؉ T cells, we analyzed patterns of DNase I hypersensitivity and epigenetic modifications within the IL-13 locus in cord blood CD4 ؉ T cells, naive or differentiated in vitro under Th1-or Th2-polarizing conditions. In naive CD4؉ T cells, hypersensitivity associated with DNA hypomethylation was limited to the distal promoter. Unexpectedly, during both Th1 and Th2 differentiation, the locus was extensively remodeled, as revealed by the formation of numerous HS sites and decreased DNA methylation. Obvious differences in chromatin architecture were limited to the proximal promoter, where strong hypersensitivity, hypomethylation, and permissive histone modifications were found selectively in Th2 cells. In addition to revealing the locations of putative cis-regulatory elements that may be required to control IL-13 expression in neonatal CD4 ؉ T cells, our results suggest that differential IL-13 expression may depend on the acquisition of a permissive chromatin architecture at the proximal promoter in Th2 cells rather than the formation of locus-wide repressive chromatin in Th1 cells.

We previously reported that the loss of E-cadherin confers a poor prognosis in lung cancer patients and is associated with in vitro resistance to EGFR inhibitors. We also demonstrated that ZEB-1 is the predominant transcriptional suppressor of E-cadherin in NSCLC cell lines. We now report that treatment with MS-275, compared to vorinostat (SAHA), valproic acid or TSA, was most effective in E-cadherin up-regulation and persistence in NSCLCs. As with other tumor types and HDAC inhibitors, MS-275 inhibited growth and induced apoptosis. Importantly, blocking E-cadherin induction by shRNA resulted in less inhibition by MS-275, implicating the EMT process as a contributing factor. In contrast to H460 and H661, H157 cells were resistant to E-cadherin up-regulation by HDAC inhibitors. This resistance was overcome, in a synergistic manner, by combined knockdown of ZEB-1 and ZEB-2. In addition, H157 cells stably transfected with E-cadherin were markedly attenuated in their tumor forming ability. Lastly, combining MS-275 with the microtubule stabilizing agent, paclitaxel, or 17-AAG, an HSP 90 inhibitor, resulted in synergistic growth inhibition. Since MS-275 has no reported activity against HDAC6, which regulates both microtubule and HSP 90 functions, other mechanisms of synergy are anticipated. These results support the role of ZEB proteins and HDAC inhibitors in the pathogenesis and treatment of lung cancer.