PRDX6, a member of the peroxiredoxins (PRDXs) family, is a key player in the removal of reactive oxygen species (ROS). Using targeted inactivation of the Prdx6 gene, we present evidence that the corresponding protein offsets the deleterious effects of ROS on lens epithelial cells (LECs) and regulates gene expression by limiting its levels. PRDX6-depleted LECs displayed phenotypic alterations and elevated a-smooth muscle actin and big-h3 expression (markers for cataractogenesis), indistinguishable from transforming growth factor b (TGFb)-induced changes. Biochemical assays disclosed enhanced levels of ROS, as well as high expression and activation of TGFb1 in Prdx6 À/À LECs. A CAT assay revealed transcriptional repression of lens epithelium-derived growth factor (LEDGF), HSP27, and aB-crystallin promoter activities in these cells. A gel mobility shift assay demonstrated the attenuation of LEDGF binding to heat shock or stress response elements present in these genes. A supply of PRDX6 toPrdx6 À/À LECs reversed these changes. Based on the above data, we propose a rheostat role for PRDX6 in regulating gene expression by controlling the ROS level to maintain cellular homeostasis.

Lens epithelium-cell derived growth factor (LEDGF) is a transcriptional activator. It protects the cells by binding to cis-stress response ((A/T)GGGG(T/A)), and heat shock (HSE; nGAAn) elements in the stress genes and activating their transcription. Transforming growth factor-␤ (TGF-␤) has been implicated in the control of tissue homeostasis, terminal differentiation, and apoptosis. Here we provide evidence that TGF-␤1 downregulates LEDGF expression and diminishes its affinity for DNA during TGF-␤1-induced phenotypic changes and apoptosis in human lens epithelial cells. Surprisingly, TGF-␤1 treatment for 48 h markedly decreased the LEDGF, Hsp27, and ␣B-crystallin promoter activities with the decrease of abundance of LEDGF mRNA and protein. Deletion mutants of the LEDGF promoter showed that one TGF-␤1 inhibitory element (TIE) like sequence nnnTTGGnnn (؊444 to ؊433) contributed to this negative regulation. Mutation of TIE (TTGG to TATT) abolished the down-regulation of the LEDGF promoter. Gel mobility and supershift assays showed that LEDGF in the nuclear extracts of TGF-␤1-treated human lens epithelial cells did not bind to stress-response elements and HSE. The TGF-␤1-induced down-regulation of LEDGF, Hsp27, and ␣B-crystallin promoters activity was reversed by cotransfection with a plasmid expressing LEDGF. Because overexpression of LEDGF was able to relieve TGF-␤1 and/or stress-induced changes, it would be a candidate molecule to postpone age-related degenerating disorders.

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