RNA interactions with CTCF are essential for its proper function

Abstract

The function of the CCCTC-binding factor (CTCF) in the organization of the genome has become an important area of investigation, but the mechanisms of how CTCF dynamically contributes to genome organization is not clear. We previously discovered that CTCF binds to large numbers of endogenous RNAs; promoting its oligomerization. Here we found that inhibition of transcription or interfering with CTCF ability to bind RNA through mutations of two of its 11 zinc fingers that are not involved with CTCF binding to its cognate site in vitro, zinc finger-1 (ZF1) or -10 (ZF10), disrupt CTCF association to chromatin. These mutations alter gene expression profiles as CTCF mutants lose their ability to promote local insulation. Our results highlight the importance of RNA as a structural component of the genome, in part by affecting the association of CTCF with chromatin and likely its interaction with other factors. Bullet Points (4 of max 75 words):• Transcriptional inhibition disrupts CTCF binding to chromatin • RNA-binding regions (RBR) in CTCF are found within ZF1 and ZF10 • Local insulation is markedly decreased in ZF1∆ and ZF10∆ mutant rescues • Gene expression and chromatin organization are disrupted by RBR mutants

PRC2 is high maintenance

“…9A,B). Given that the stability of CTCF on chromatin requires its RNA binding activity (Hansen et al 2018;Saldana-Meyer et al 2019), we suspect that disrupting this latter activity would give rise to similar developmental defects.…”

CTCF controls imprinted gene activity at the mouseDlk1-Dio3andIgf2-H19domains by modulating allele-specific sub-TAD structure

“…Recently, we found that the maternal expression of Meg3, and possibly the lncRNA itself, prevents activation of Dlk1 as well (25). We hypothesise that the two mechanisms are linked, with either the Dlk1-Meg3 sub-TAD focusing or constraining the repressive function of the Meg3 expression or transcript, or conversely, with maternal Meg3 expression facilitating CTCF recruitment or its stability of binding at the DMR, possibly through direct RNA-CTCF contacts (32,33).…”

Functional conservation of lncRNAJPXdespite sequence and structural divergence

“…In addition, we also found human lncRNA JPX present as one of the RNA transcripts pulled-down with human CTCF by PAR-CLIP from human bone osteosarcoma U2OS cells (Supplementary data from (Saldaña-Meyer et al, 2014)). Moreover, RNA-binding regions (RBR) in CTCF have been recently reported and shown to be essential for the molecular interaction and function of CTCF in mouse and human cells (Hansen et al, 2018;Saldana-Meyer et al, 2019), which confirm CTCF binding to endogenous RNAs and the functional importance of CTCF-RNA binding in gene regulation. Our evidence of direct binding of CTCF to human JPX RNA similar to CTCF-Jpx lncRNA interaction in the mouse thus supports the functional importance and conservation of human JPX to its mouse homolog.…”