Induction of anti-aging gene klotho with a small chemical compound that demethylates CpG Islands

Klotho (KL) is described as an anti-aging gene because mutation of KI gene leads to multiple pre-mature aging phenotypes and shortens lifespan in mice. Growing evidence suggests that an increase in KL expression may be beneficial for agerelated diseases such as arteriosclerosis and diabetes. It remains largely unknown, however, how KI expression could be induced. Here we discovered novel molecular mechanism for induction of KI expression with a small molecule 'Compound H', N-(2-chlorophenyl)-1H-indole-3-caboxamide. Compound H was originally identified through a high-throughput screening of small molecules for identifying KI inducers. However, how Compound H induces KI expression has never been investigated. We found that Compound H increased KI expression via demethylation in CpG islands of the KI gene. The demethylation was accomplished by activating demethylases rather than inhibiting methylases. Due to demethylation, Compound H enhanced binding of transcription factors, Pax4 and Kid3, to the promoter of the KI gene. Pax4 and Kid3 regulated KI promoter activity positively and negatively, respectively. Thus, our results show that demethylation is an important molecular mechanism that mediates Compound H-induced KI expression. Further investigation is warranted to determine whether Compound H demethylates the KI gene in vivo and whether it can serve as a therapeutic agent for repressing or delaying the onset of age-related diseases.