Multivalent Histone and DNA Engagement by a PHD/BRD/PWWP Triple Reader Cassette Recruits ZMYND8 to K14ac-Rich Chromatin

摘要

class="head no_bottom_margin" id="sec1title">IntroductionPost-translational modifications (PTMs) on histones and DNA are critical regulators of chromatin stability, structure, and gene expression (, ). Combinations of histone PTMs have been recognized to constitute a cellular language, involving deposition, interpretation, and removal of PTMs, referred to as the “histone code” (). Different classes of protein interaction modules (“readers”) have evolved to recognize and bind to specific PTMs, including lysine methylation and acetylation, among many known PTMs (, ). Acetylated lysines (Kac) are recognized by bromodomain modules (BRDs) with affinities ranging in the micromolar range (); a single BRD can also bind two adjacent acetylation marks (, ), thus enhancing affinity. Recognition of multiple histone PTMs by employing several effector modules present within a given protein confers avidity and increases specificity/affinity as a result of multivalency ().Multivalency has been studied in BRD-containing proteins involving two effector modules, often another BRD or a plant homeodomain (PHD). For example, in the case of the transcription initiation factor TFIID subunit 1 (TAF1), two BRD modules exist in a rigidly confined topology, allowing for optimal recognition of multiple acetylated lysines on histone H4 with low micromolar affinity (). Multi-domain or paired arrangements including a PHD/BRD cassette provide a platform for combinatorial recognition of lysine methylation and acetylation, introducing higher specificity. For example, the chromatin regulator tripartite motif 24 (TRIM24) binds to the N-terminal H3 tail, simultaneously engaging lysine 4 (H3K4), via a PHD finger, whereas an adjacent BRD module linked via a flexible loop binds to acetylated lysine 23 (H3K23ac) (). The relative topology between two effector modules affects the type of signals interpreted as a function of the linker connecting the modules that acts as a molecular ruler; for instance, the PHD/BRD cassette found in BPTF can recognize and bind to histone 3 dimethyl-lysine 4 (H3K4me2) and histone 3 trimethyl-lysine 4 (H3K4me3) () via the PHD domain, facilitating the adjacent BRD to gain specificity for histone 4 lysine 16 acetyl (H4K16ac) found in trans within a single nucleosome over other H4 acetylations (). Therefore, combination of effector modules not only introduces conformational plasticity, leading to specificity, but also introduces avidity, resulting in higher affinities. However, it is not clear whether this is an additive effect so that introduction of additional modules will result in similar enhancements offering specificity toward developing landscapes of PTMs.To better understand how multivalent interactions involving more than two reader domains impact recognition of PTMs, affecting protein function, we studied the topology of the N-terminal triple reader domain architecture found in the zinc-finger MYND domain-containing protein 8 (ZMYND8), which contains, in addition to a PHD/BRD arrangement, a Pro-Trp-Trp-Pro (PWWP) domain within a PHD/BRD/PWWP cassette. ZMYND8 has been previously found to participate in transcriptional regulation complexes (, , href="#bib16" rid="bib16" class=" bibr popnode">Kloet et al., 2015, href="#bib21" rid="bib21" class=" bibr popnode">Malovannaya et al., 2011), is implicated in gene silencing (href="#bib28" rid="bib28" class=" bibr popnode">Poleshko et al., 2010), acts as a DNA damage response element (href="#bib10" rid="bib10" class=" bibr popnode">Gong et al., 2015), and has recently been proposed to control, together with KDM5C, enhancer activity (href="#bib36" rid="bib36" class=" bibr popnode">Shen et al., 2016). Here we describe the high-resolution crystal structure of the ZMYND8 N-terminal triple reader PHD/BRD/PWWP module and show how contributions from multivalent, simultaneous recognition of DNA and histone PTMs drive ZMYND8 function, affecting recruitment to DNA-damaged sites.