BZLF1 interacts with chromatin remodelers promoting escape from latent infections with EBV.

Marisa Schaeffner,Anne Woellmer,Wolfgang Hammerschmidt,Paulina Mrozek-Gorska,Takanobu Tagawa,Alexander Buschle,Filippo M Cernilogar,Corinna Lieleg,Nils Krietenstein,Philipp Korber,Gunnar Schotta

doi:10.26508/lsa.201800108

Abstract

A hallmark of EBV infections is its latent phase, when all viral lytic genes are repressed. Repression results from a high nucleosome occupancy and epigenetic silencing by cellular factors such as the Polycomb repressive complex 2 (PRC2) and DNA methyltransferases that, respectively, introduce repressive histone marks and DNA methylation. The viral transcription factor BZLF1 acts as a molecular switch to induce transition from the latent to the lytic or productive phase of EBV's life cycle. It is unknown how BZLF1 can bind to the epigenetically silenced viral DNA and whether it directly reactivates the viral genome through chromatin remodeling. We addressed these fundamental questions and found that BZLF1 binds to nucleosomal DNA motifs both in vivo and in vitro. BZLF1 co-precipitates with cellular chromatin remodeler ATPases, and the knock-down of one of them, INO80, impaired lytic reactivation and virus synthesis. In Assay for Transposase-Accessible Chromatin-seq experiments, non-accessible chromatin opens up locally when BZLF1 binds to its cognate sequence motifs in viral DNA. We conclude that BZLF1 reactivates the EBV genome by directly binding to silenced chromatin and recruiting cellular chromatin-remodeling enzymes, which implement a permissive state for lytic viral transcription. BZLF1 shares this mode of action with a limited number of cellular pioneer factors, which are instrumental in transcriptional activation, differentiation, and reprogramming in all eukaryotic cells.

Highlights

Eukaryotic DNA-binding sites are often not accessible to their cognate factors because the sites lie within epigenetically silent chromatin and are occupied by nucleosomes
We found that the BZLF1 levels we reach in the Raji inducible system are in a range found in the small fraction of B95-8 cells that undergo the lytic cycle of EBV (Buschle et al, 2019 Preprint)
The BZLF1 basic leucinezipper (bZIP) domain alone, that is, BZLF1 lacking its transactivation domain (TAD) and the ultimate carboxy terminus, interacted with SNF2h but not with INO80 (Fig 5A). These results suggested that BZLF1 interacts with subunits of at least two cellular chromatin remodeler families possibly to recruit them to lytic gene promoters and support EBV’s lytic reactivation

Summary

Introduction

Eukaryotic DNA-binding sites are often not accessible to their cognate factors because the sites lie within epigenetically silent chromatin and are occupied by nucleosomes. Pioneer factors either open chromatin directly through their binding or recruit chromatin modifiers and ATP-dependent chromatin-remodeling enzymes that open chromatin to allow access for the transcription machinery (Clapier & Cairns, 2009; Bartholomew, 2014; Langst & Manelyte, 2015). Such pioneer factors play key roles in hormone-dependent cancers (Jozwik & Carroll, 2012), embryonic stem cells and cell fate specification (Smale, 2010; Drouin, 2014), and cellular reprogramming (Iwafuchi-Doi & Zaret, 2014; Soufi et al, 2015). 2,000–3,000 sequencespecific DNA-binding transcription factors in human cells are known (Lander et al, 2001; Venter et al, 2001), but only about a dozen are functionally confirmed as pioneer factors

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Conclusion