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

Dongju Jung,Yuechi Xu,Zhongjie Sun

doi:10.18632/oncotarget.18608

Dongju Jung, Yuechi Xu + Show 1 more

Open Access

https://doi.org/10.18632/oncotarget.18608

Copy DOI

Abstract

Klotho (KL) is described as an anti-aging gene because mutation of Kl 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 age-related diseases such as arteriosclerosis and diabetes. It remains largely unknown, however, how Kl expression could be induced. Here we discovered novel molecular mechanism for induction of Kl 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 Kl inducers. However, how Compound H induces Kl expression has never been investigated. We found that Compound H increased Kl expression via demethylation in CpG islands of the Kl 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 Kl gene. Pax4 and Kid3 regulated Kl promoter activity positively and negatively, respectively. Thus, our results show that demethylation is an important molecular mechanism that mediates Compound H-induced Kl expression. Further investigation is warranted to determine whether Compound H demethylates the Kl gene in vivo and whether it can serve as a therapeutic agent for repressing or delaying the onset of age-related diseases.

Highlights

Pre-mature aging phenotypes were eminent in the klotho (Kl)-deficient mice, which have ~ 10 copies of a transgene integrated in the 5’ flanking region of the Kl gene disrupting its expression [1]
This study demonstrates a new molecular mechanism for induction of Kl expression, i.e., demethylation of Kl gene, by a small molecule called compound H
We found that compound H activated the expression of a reporter plasmid, which was composed of human KL gene promoter followed by luciferase gene [21]

Summary

INTRODUCTION

Pre-mature aging phenotypes were eminent in the klotho (Kl)-deficient mice, which have ~ 10 copies of a transgene integrated in the 5’ flanking region of the Kl gene disrupting its expression [1]. Two other Kl-related genes have been identified based on the DNA sequence similarity; they are designated as β- and γ-klotho, respectively [12] They are type I transmembrane proteins and behave as coreceptors of FGFs similar to the original Kl which is designated as αKl, but they are expressed in different tissues [13]. In an effort to find molecular mechanisms for Kl induction, we examined the small molecules and transcription factors that bind to the promoter/enhancer sequences of Kl gene. Transactivation functions of the selected transcription factors and small molecules were investigated using a reporter plasmid in which firefly luciferase gene expression was under control of human KL promoter/enhancer [21].We found an activation mechanism for induction of Kl expression by a small chemical compound H. DNA demethylation could be an activation mechanism for Kl expression

RESULTS

DISCUSSION

MATERIALS AND METHODS