Francesco Gualdrini2, Sara Polletti2, Marta Simonatto1, Elena Prosperini, Francesco Pileri and Gioacchino Natoli European Institute of Oncology (IEO), Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS), Milan 20139, Italy Corresponding authors: gioacchino.natoliieo.it, francesco.gualdriniieo.it, sara.pollettiieo.it

↵2 These authors contributed equally to this work.

↵1 Present address: Directorate of Research Services and Valorization, University of Trento, Trento 38122, Italy.

Abstract

Six methyltransferases divide labor in establishing genomic profiles of histone H3 lysine 9 methylation (H3K9me), an epigenomic modification controlling constitutive heterochromatin, gene repression, and silencing of retroelements. Among them, SETDB1 is recruited to active chromatin domains to silence the expression of endogenous retroviruses. In the context of experiments aimed at determining the impact of SETDB1 on stimulus-inducible gene expression in macrophages, we found that loss of H3K9me3 caused by SETDB1 depletion was associated with increased recruitment of CTCF to >1600 DNA binding motifs contained within SINE B2 repeats, a previously unidentified target of SETDB1-mediated repression. CTCF is an essential regulator of chromatin folding that restrains DNA looping by cohesin, thus creating boundaries among adjacent topological domains. Increased CTCF binding to SINE B2 repeats enhanced insulation at hundreds of sites and increased loop formation within topological domains containing lipopolysaccharide-inducible genes, which correlated with their impaired regulation in response to stimulation. These data indicate a role of H3K9me3 in restraining genomic distribution and activity of CTCF, with an impact on chromatin organization and gene regulation.

Received December 7, 2021. Accepted March 16, 2022.