Crystal structures of REF6 and its complex with DNA reveal diverse recognition mechanisms

Zizi Tian,Zhihui Li,Hon-Ming Lam,Cody Caba,Xiaorong Li,Lulu Ma,Manfeng Zhang,Zhenhang Chen,Shaodong Dai,Meiting Yang,Chenjun Tang,Min Li,Zhongzhou Chen,Yufeng Tong,Yunlong Liu,Wei Wu

doi:10.1038/s41421-020-0150-6

Zizi Tian, Zhihui Li + Show 14 more

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https://doi.org/10.1038/s41421-020-0150-6

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Relative of Early Flowing 6 (REF6) is a DNA-sequence-specific H3K27me3/2 demethylase that contains four zinc finger (ZnF) domains and targets several thousand genes in Arabidopsis thaliana. The ZnF domains are essential for binding target genes, but the structural basis remains unclear. Here, we determined crystal structures of the ZnF domains and REF6-DNA complex, revealing a unique REF6-family-specific half-cross-braced ZnF (RCZ) domain and two C2H2-type ZnFs. DNA-binding induces a profound conformational change in the hinge region of REF6. Each REF6 recognizes six bases and DNA methylation reduces the binding affinity. Both the acidic region and basic region are important for the self-association of REF6. The REF6 DNA-binding affinity is determined by the sequence-dependent conformations of DNA and also the cooperativity in different target motifs. The conformational plasticity enables REF6 to function as a global transcriptional regulator that directly binds to many diverse genes, revealing the structural basis for the epigenetic modification recognition.

Highlights

Histones are subjected to comprehensive posttranslational modifications, including methylation, acetylation, phosphorylation, and ubiquitination
We identified a novel Relative of Early Flowering 6 (REF6)-family-specific half-cross-braced zinc finger (ZnF) (RCZ) domain, a hinge, an acidic region, a basic region, and two classical C2H2 ZnF domains in the structure
The ZnF domains found in histone demethylases are primarily involved in binding histones or ubiquitination

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Introduction

Histones are subjected to comprehensive posttranslational modifications, including methylation, acetylation, phosphorylation, and ubiquitination. Histone methylation generally occurs on lysine and arginine residues of the N-terminal tails and this can provide an activating or repressive epigenetic mark. No homologs of metazoan H3K27-specific demethylases KDM6a or KDM6b were found in Arabidopsis[5]. Both Relative of Early Flowering 6 (REF6)[6] and Early Flowering 6 (ELF6)[7] have been shown to be specific H3K27me3/2 demethylases in plants. JMJ13 is an H3K27me[3] site-specific demethylase, but has no significant activity on H3K27me[28]. REF6, ELF6 and JMJ13 are homologous and belong to the KDM4 subfamily

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