N⁶-methyladenosine (m⁶A) is the most abundant internal RNA modification of cellular mRNAs. m⁶A is recognised by YTH domain-containing proteins, which selectively bind to m⁶A-decorated RNAs regulating their turnover and translation. Using an m⁶A-modified hairpin present in the Kaposi’s sarcoma associated herpesvirus (KSHV) ORF50 RNA, we identified seven members from the ‘Royal family’ as putative m⁶A readers, including SND1. RIP-seq and eCLIP analysis characterised the SND1 binding profile transcriptome-wide, revealing SND1 as an m⁶A reader. We further demonstrate that the m⁶A modification of the ORF50 RNA is critical for SND1 binding, which in turn stabilises the ORF50 transcript. Importantly, SND1 depletion leads to inhibition of KSHV early gene expression showing that SND1 is essential for KSHV lytic replication. This work demonstrates that members of the ‘Royal family’ have m⁶A-reading ability, greatly increasing their epigenetic functions beyond protein methylation.

Reactivation of lytic replication from viral latency is a defining property of all herpesviruses. Despite this, the authentic physiological cues for the latent-lytic switch are unclear. Such cues should ensure that viral lytic replication occurs under physiological conditions, predominantly in sites which facilitate transmission to permissive uninfected cells and new susceptible hosts. Kaposi's sarcoma-associated herpesvirus (KSHV) is associated with the B-cell neoplasm primary effusion lymphoma (PEL), in which the virus remains latent. We have previously shown that PEL cells have the gene expression profile and immunophenotype of cycling preplasma cells (plasmablasts). Here, we show that the highly active spliced isoform of plasma cell transcription factor X box binding protein 1 (XBP-1s) is a lytic switch for KSHV. XBP-1s is normally absent in PEL, but the induction of endoplasmic reticulum stress leads to XBP-1s generation, plasma cell-like differentiation, and lytic reactivation of KSHV. XBP-1s binds to and activates the KSHV immediate-early gene ORF50 and synergizes with the ORF50 gene product RTA to induce a full lytic cycle. These data suggest that KSHV remains latent until B-cell terminal differentiation into plasma cells, the transcriptional environment of which provides the physiological “lytic switch” through XBP-1s. This links B-cell terminal differentiation to KSHV lytic reactivation.

Kaposi's Sarcoma-associated herpesvirus (KSHV) and Epstein Barr virus (EBV) are the causative agents of several malignancies. Like all herpesviruses, KSHV and EBV undergo distinct latent and lytic replication programmes. The transition between these states allows the establishment of a lifelong persistent infection, dissemination to sites of disease and the spread to new hosts. Latency-associated viral proteins have been well characterised in transformation and tumourigenesis pathways; however, a number of studies have shown that abrogation of KSHV and EBV lytic gene expression impairs the oncogenesis of several cancers. Furthermore, several lytically expressed proteins have been functionally tethered to the angioproliferative and anti-apoptotic phenotypes of virus-infected cells. As a result, the investigation and therapeutic targeting of KSHV and EBV lytic cycles may be essential for the treatment of their associated malignancies.

The coronavirus nucleocapsid (N) protein plays a multifunctional role in the virus life cycle, from regulation of replication and transcription and genome packaging to modulation of host cell processes. These functions are likely to be facilitated by interactions with host cell proteins. The potential interactome of the infectious bronchitis virus (IBV) N protein was mapped using stable isotope labeling with amino acids in cell culture (SILAC) coupled to a green fluorescent protein-nanotrap pulldown methodology and liquid chromatography-tandem mass spectrometry. The addition of the SILAC label allowed discrimination of proteins that were likely to specifically bind to the N protein over background binding. Overall, 142 cellular proteins were selected as potentially binding to the N protein, many as part of larger possible complexes. These included ribosomal proteins, nucleolar proteins, translation initiation factors, helicases, and hnRNPs. The association of selected cellular proteins with IBV N protein was confirmed by immunoblotting, cosedimentation, and confocal microscopy. Further, the localization of selected proteins in IBV-infected cells as well as their activity during virus infection was assessed by small interfering RNA-mediated depletion, demonstrating the functional importance of cellular proteins in the biology of IBV. This interactome not only confirms previous observations made with other coronavirus and IBV N proteins with both overexpressed proteins and infectious virus but also provides novel data that can be exploited to understand the interaction between the virus and the host cell.

Merkel cell carcinoma (MCC) is a highly aggressive nonmelanoma skin cancer arising from epidermal mechanoreceptor Merkel cells. In 2008, a novel human polyomavirus, Merkel cell polyomavirus (MCPyV), was identified and is strongly implicated in MCC pathogenesis. Currently, little is known regarding the virus-host cell interactions which support virus replication and virus-induced mechanisms in cellular transformation and metastasis. Here we identify a new function of MCPyV small T antigen (ST) as an inhibitor of NF-κB-mediated transcription. This effect is due to an interaction between MCPyV ST and the NF-κB essential modulator (NEMO) adaptor protein. MCPyV ST expression inhibits IκB kinase α (IKKα)/IKKβ-mediated IκB phosphorylation, which limits translocation of the NF-κB heterodimer to the nucleus. Regulation of this process involves a previously undescribed interaction between MCPyV ST and the cellular phosphatase subunits, protein phosphatase 4C (PP4C) and/or protein phosphatase 2A (PP2A) Aβ, but not PP2A Aα. Together, these results highlight a novel function of MCPyV ST to subvert the innate immune response, allowing establishment of early or persistent infection within the host cell.