Optogenetic regulation of site-specific subtelomeric DNA-methylation

// Samrat Roy Choudhury 1 , Yi Cui 1 , Anoop Narayanan 2 , David P. Gilley 3 , Nazmul Huda 3 , Chiao-Ling Lo 4 , Feng C. Zhou 4, 5 , Dinesh Yernool 2 , Joseph Irudayaraj 1 1 Department of Agricultural & Biological Engineering, Bindley Bioscience Center, Purdue Center for Cancer Research, Purdue University, West Lafayette, IN 47907, USA 2 Bindley Laboratory of Structural Biology, Bindley Bioscience Center, Purdue University, West Lafayette, IN 47907, USA 3 Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, IN 46202, USA 4 Department of Anatomy and Cell Biology, Indiana University School of Medicine, Indianapolis, IN 46202, USA 5 Stark Institute of Neuroscience Research, Indiana University School of Medicine, Indianapolis, IN 46202, USA Correspondence to: Dinesh Yernool, email: dyernool@purdue.edu Joseph Irudayaraj, email: josephi@purdue.edu Keywords: optogenetics, single-cells tools, subtelomeric DNA-methylation, telomere-elongation Received: April 25, 2016      Accepted: June 18, 2016      Published: July 04, 2016 ABSTRACT Telomere length homeostasis, critical for chromosomal integrity and genome stability, is controlled by intricate molecular regulatory machinery that includes epigenetic modifications. Here, we examine site-specific and spatiotemporal alteration of the subtelomeric methylation of CpG islands using optogenetic tools to understand the epigenetic regulatory mechanisms of telomere length maintenance. Human DNA methyltransferase3A (DNMT3A) were assembled selectively at chromosome ends by fusion to cryptochrome 2 protein (CRY2) and its interacting complement, the basic helix loop helix protein-1 (CIB1). CIB1 was fused to the telomere-associated protein telomere repeat binding factor-1 (TRF1), which localized the protein complex DNMT3A-CRY2 at telomeric regions upon excitation by blue-light monitored by single-molecule fluorescence analyses. Increased methylation was achieved selectively at subtelomeric CpG sites on the six examined chromosome ends specifically after blue-light activation, which resulted in progressive increase in telomere length over three generations of HeLa cell replications. The modular design of the fusion constructs presented here allows for the selective substitution of other chromatin modifying enzymes and for loci-specific targeting to regulate the epigenetic pathways at telomeres and other selected genomic regions of interest.