Pronounced sequence specificity of the TET enzyme catalytic domain guides its cellular function
Mirunalini Ravichandran, Dominik Rafalski, Claudia I. Davies, Oscar Ortega‐Recalde, Xinsheng Nan, Cassandra R. Glanfield, Annika Kötter, Katarzyna Misztal, Andrew H. Wang, Marek Wojciechowski, Michał Rażew, Issam M. Mayyas, Olga Kardailsky, Uwe Schwartz, Krzysztof Zembrzycki, Ian M. Morison, Mark Helm, Dieter Weichenhan, Renata Z. Jurkowska, Felix Krueger, Christoph Plass, Martin Zacharias, Matthias Bochtler, Timothy A. Hore, Tomasz P. Jurkowski
Abstract
TET (ten-eleven translocation) enzymes catalyze the oxidation of 5-methylcytosine bases in DNA, thus driving active and passive DNA demethylation. Here, we report that the catalytic domain of mammalian TET enzymes favor CGs embedded within basic helix-loop-helix and basic leucine zipper domain transcription factor-binding sites, with up to 250-fold preference in vitro. Crystal structures and molecular dynamics calculations show that sequence preference is caused by intrasubstrate interactions and CG flanking sequence indirectly affecting enzyme conformation. TET sequence preferences are physiologically relevant as they explain the rates of DNA demethylation in TET-rescue experiments in culture and in vivo within the zygote and germ line. Most and least favorable TET motifs represent DNA sites that are bound by methylation-sensitive immediate-early transcription factors and octamer-binding transcription factor 4 (OCT4), respectively, illuminating TET function in transcriptional responses and pluripotency support.