Brd/BET Proteins Influence the Genome-Wide Localization of the Kaposi’s Sarcoma-Associated Herpesvirus and Murine Gammaherpesvirus Major Latency Proteins

Abstract

The rhadinoviruses Kaposi's Sarcoma-associated herpesvirus (KSHV) and murine gammaherpesvirus (MHV-68) persist in infected hosts in a latent state that is characterized by the absence of virus production and by restricted viral gene expression. Their major latency protein, the latency-associated nuclear antigen (kLANA for KSHV and mLANA for MHV-68), is essential for viral genome maintenance and replication and involved in transcriptional regulation. Both kLANA and mLANA interact with cellular chromatin-associated proteins, among them the Bromodomain and Extra Terminal domain (Brd/BET) proteins, which recruit cellular and viral proteins to acetylated histones through their bromodomains and modulate cellular gene expression. Brd/BET proteins also play a role in the tethering, replication, segregation or integration of a diverse group of viral DNA genomes. In this study we explored if Brd/BET proteins influence the localization of the LANAs to preferential regions in the host chromatin and thereby contribute to kLANA-or mLANA-mediated transcriptional regulation. Using ChIP-Seq, we revealed a genome-wide co-enrichment of kLANA with Brd2/4 near cellular and viral transcriptional start sites (TSS). Treatment with I-BET151, an inhibitor of Brd/BET, displaced kLANA and Brd2/4 from TSS in the viral and host chromatin, but did not affect the direct binding of kLANA to kLANA-binding sites (LBS) in the KSHV latent origin of replication. Similarly, mLANA, but not a mLANA mutant deficient for binding to Brd2/4, also associated with cellular TSS. We compared the transcriptome of KSHV-infected with uninfected and kLANA-expressing human B cell lines, as well as a murine B cell line expressing mLANA or a Brd2/4-binding deficient mLANA mutant. We found that only a minority of cellular genes, whose TSS are occupied by kLANA or mLANA, is transcriptionally regulated by these latency proteins. Our findings extend previous reports on a preferential deposition of kLANA on cellular TSS and show that this characteristic chromatin association pattern is at least partially determined by the interaction of these viral latency proteins with members of the Brd/BET family of chromatin modulators.

Relevance of BET Family Proteins in SARS-CoV-2 Infection

“…In this work, authors have shown that treatment with the BET inhibitor I-BET151 displaces kLANA protein, as well as BRD2 and BRD4, from viral and host TSSs. Mutations in mLANA preventing BRD2 and BRD4 binding [158] also have similar consequences [98]. In turn, EBNA1 has been described to interact with BRD4 and this interaction appears to play a critical role in EBNA1-mediated transcriptional activation, as it has been shown that BRD4 silencing leads to a decrease in its transcriptional activity [100].…”

“…Its homologue in MHV-68, mLANA, is expressed in latency and during lytic replication and its function is essential in the establishment and maintenance of latency [147][148][149][150][151][152] and the EBV homologue, the Epstein-Barr virus nuclear antigen 1 (EBNA1), is involved in the regulation of viral transcription, replication and persistence [153]. Several studies show that kLANA and mLANA interact with a region containing the ET domain of BRD2, BRD3 and BRD4 [94][95][96][97]99], and a recent study has described that the preferential localization of kLANA and mLANA at transcription start sites (TSSs) [154][155][156][157] is due in part to BET proteins [98]. In this work, authors have shown that treatment with the BET inhibitor I-BET151 displaces kLANA protein, as well as BRD2 and BRD4, from viral and host TSSs.…”

Repurposing clinically available drugs and therapies for pathogenic targets to combat SARS‐CoV‐2

“… 203 BRD2 is closely linked to infection by various types of viruses, including Kaposi's sarcoma associated‐herpesvirus, murine herpesvirus68, Epstein–Barr virus, and murine leukemia virus. 204 , 205 , 206 , 207 , 208 , 209 Since the infection of host cells by COVID‐19 mainly depends on the RBD domain of the Spike protein, researchers used RBD binding as a phenotypic readout in a high‐throughput CRISPRi screen to systematically discover significant factors regulating viral infection in host cells. 167 The results demonstrated that knocking out the BRD2 gene effectively prevented RBD binding.…”

Viral Manipulation of the Host Epigenome as a Driver of Virus-Induced Oncogenesis

“…Even though the binding sites identified in these studies may partially differ, due to the use of a different cellular systems, they show that LANA binds to a far larger number of sites than previously thought. Moreover, binding is preferentially targeted to euchromatic loci, possibly due to LANA's interactions with the H3K4 methyltransferase hSET1 or with BET [253][254][255][256]. It is also of interest that the expression of genes in the proximity of those binding sites does not seem to be affected, suggesting that methylation mediated silencing may be context specific.…”