Structural insights into H2A-H2B and H2A.Z-H2B sliding on histone chaperone NAP1

Abstract

The evolutionarily conserved nucleosome assembly protein 1 (NAP1) functions as a histone chaperone for H2A-H2B, regulating nucleosome assembly and maintaining chromatin integrity. However, the dynamic and variable nature of the interactions between acidic NAP1 and basic H2A-H2B has obscured the molecular basis of its chaperoning activity. Here, we report the crystal structures of Caenorhabditis elegans NAP1 (CeNAP1) in complex with Xenopus laevis H2A-H2B (XlH2A-H2B) and with C. elegans H2A.Z-H2B (CeH2A.Z-H2B) at 3.35 Å and 2.8 Å, respectively. In our structures, H2A/H2A.Z-H2B binds to the acidic concave surface of CeNAP1 in three distinct poses, with two in the CeNAP1-XlH2A-H2B complex and one in the CeNAP1-CeH2A.Z-H2B complex. These poses are different from the two poses observed in the previously reported CeNAP1-CeH2A/H2A.Z-H2B structures. The predominant interaction involves engagement of the acidic CeNAP1 α6-carboxy-terminal (C-terminal) tail by the basic H2A/H2A.Z αN–α1 region, stabilized by salt bridges and electrostatic interactions. A comparative analysis of all five known poses reveals that H2A/H2A.Z-H2B can shift approximately 20.7 Å along the α6-C-terminal tail-C′-terminal tail-α6′ axis. These findings demonstrate a sliding binding mode of H2A/H2A.Z-H2B on NAP1, providing new mechanistic insights into nucleosome assembly activity of histone chaperones.

1. Introduction

In eukaryotes, genetic information is packaged into chromatin, whose fundamental structural unit is the nucleosome. Each nucleosome is composed of a histone octamer, formed by two H2A-H2B dimers and an H3-H4 tetramer, wrapped with ~147 base pairs of DNA [1]. The dynamic disassembly and reassembly of this structure are integral to essential genomic processes such as DNA replication, transcription, and damage repair [2–4], making precise nucleosome dynamics critical for maintaining genome integrity [5–7]. This dynamic is facilitated by histone chaperones, a large family of proteins that bind to H2A-H2B or H3-H4 [8] to escort their orderly deposition onto or eviction from acidic DNA, thereby preventing aberrant non-specific interactions [9].

As a major histone chaperone, nucleosome assembly protein 1 (NAP1), specially handles canonical H2A-H2B and the variant H2A.Z-H2B, playing an essential role in their incorporation into chromatin [10–13]. The chaperone function of NAP1 relies on its acidic character, which allows it to compete with the similarly acidic DNA for binding to the basic surfaces of H2A-H2B dimers, effectively shielding the histones [9,14]. The electrostatic nature of this interaction is evidenced by its sensitivity to high-salt conditions [15]. Structurally, complexes of Caenorhabditis elegans NAP1 (CeNAP1) with Caenorhabditis elegans H2A-H2B (CeH2A-H2B) or C. elegans H2A.Z-H2B (CeH2A.Z-H2B) have revealed that the acidic α6 and C-terminal tail of CeNAP1 engage the basic αN-α1 region of H2A and H2A.Z through salt bridges and electrostatic interactions, mimicking the histone interaction with DNA in the nucleosome [15–17]. Notably, the presence of an extensive acidic surface presented by CeNAP1 suggests a potential that H2A–H2B can adopt multiple poses when bound to CeNAP1.

This structural plasticity implies that the precise mechanism by which NAP1 orchestrates nucleosome assembly is not fully elucidated. In this study, we investigate whether H2A-H2B can slide along CeNAP1’s acidic surface in a manner analogous to histone octamer sliding on DNA during chromatin remodeling [18]. Through structural determination and analyzation of CeNAP1-H2A/H2A.Z-H2B complexes, we reveal that H2A/H2A.Z-H2B binds to CeNAP1 in at least five distinct poses, providing direct structural evidence for a sliding binding mode. This discovery offers a new mechanistic perspective on the chaperone activity of NAP1 and its role in nucleosome assembly.