Advertising to Early Trend Propagators: Evidence from Twitter

In the digital economy, influencing and controlling the spread of information is a key concern for firms. One way firms try to achieve this is to target firm communications to consumers who embrace and propagate the spread of new information on emerging and `trending' topics on social media. However, little is known about whether early trend propagators are indeed responsive to firm-sponsored messages. To explore whether early propagators of trending topics respond to advertising messages, we use data from two field tests conducted by a charity and an emerging fashion firm on the micro-blogging service Twitter. On Twitter, 'promoted tweets' allow advertisers to target individuals based on the content of their recent postings. Twitter continuously identifies in real time which topics are newly popular among Twitter users. In the field tests, we collaborated with a charity and a fashion firm to target ads at consumers who embraced a Twitter trend early in its life-cycle by posting about it, and compared their behavior to that of consumers who posted about the same topic only later on. Throughout both field tests, we consistently find that early propagators of trends are less responsive to advertising than consumers who embrace trends later

The UFM1 Pathway Impacts HCMV US2-Mediated Degradation of HLA Class I

To prevent accumulation of misfolded proteins in the endoplasmic reticulum, chaperones perform quality control on newly translated proteins and redirect misfolded proteins to the cytosol for degradation by the ubiquitin-proteasome system. This pathway is called ER-associated protein degradation (ERAD). The human cytomegalovirus protein US2 induces accelerated ERAD of HLA class I molecules to prevent immune recognition of infected cells by CD8(+) T cells. Using US2-mediated HLA-I degradation as a model for ERAD, we performed a genome-wide CRISPR/Cas9 library screen to identify novel cellular factors associated with ERAD. Besides the identification of known players such as TRC8, p97, and UBE2G2, the ubiquitin-fold modifier1 (UFM1) pathway was found to affect degradation of HLA-I. UFMylation is a post-translational modification resembling ubiquitination. Whereas we observe ubiquitination of HLA-I, no UFMylation was detected on HLA-I or several other proteins involved in degradation of HLA-I, suggesting that the UFM1 pathway impacts ERAD in a different manner than ubiquitin. Interference with the UFM1 pathway seems to specifically inhibit the ER-to-cytosol dislocation of HLA-I. In the absence of detectable UFMylation of HLA-I, UFM1 may contribute to US2-mediated HLA-I degradation by misdirecting protein sorting indirectly. Mass spectrometry analysis of US2-expressing cells showed that ribosomal proteins are a major class of proteins undergoing extensive UFMylation; the role of these changes in protein degradation may be indirect and remains to be established

Varicellovirus UL 49.5 proteins differentially affect the function of the transporter associated with antigen processing, TAP

Cytotoxic T-lymphocytes play an important role in the protection against viral infections, which they detect through the recognition of virus-derived peptides, presented in the context of MHC class I molecules at the surface of the infected cell. The transporter associated with antigen processing (TAP) plays an essential role in MHC class I–restricted antigen presentation, as TAP imports peptides into the ER, where peptide loading of MHC class I molecules takes place. In this study, the UL49.5 proteins of the varicelloviruses bovine herpesvirus 1 (BHV-1), pseudorabies virus (PRV), and equine herpesvirus 1 and 4 (EHV-1 and EHV-4) are characterized as members of a novel class of viral immune evasion proteins. These UL49.5 proteins interfere with MHC class I antigen presentation by blocking the supply of antigenic peptides through inhibition of TAP. BHV-1, PRV, and EHV-1 recombinant viruses lacking UL49.5 no longer interfere with peptide transport. Combined with the observation that the individually expressed UL49.5 proteins block TAP as well, these data indicate that UL49.5 is the viral factor that is both necessary and sufficient to abolish TAP function during productive infection by these viruses. The mechanisms through which the UL49.5 proteins of BHV-1, PRV, EHV-1, and EHV-4 block TAP exhibit surprising diversity. BHV-1 UL49.5 targets TAP for proteasomal degradation, whereas EHV-1 and EHV-4 UL49.5 interfere with the binding of ATP to TAP. In contrast, TAP stability and ATP recruitment are not affected by PRV UL49.5, although it has the capacity to arrest the peptide transporter in a translocation-incompetent state, a property shared with the BHV-1 and EHV-1 UL49.5. Taken together, these results classify the UL49.5 gene products of BHV-1, PRV, EHV-1, and EHV-4 as members of a novel family of viral immune evasion proteins, inhibiting TAP through a variety of mechanisms

The Epstein-Barr Virus G-Protein-Coupled Receptor Contributes to Immune Evasion by Targeting MHC Class I Molecules for Degradation

Epstein-Barr virus (EBV) is a human herpesvirus that persists as a largely subclinical infection in the vast majority of adults worldwide. Recent evidence indicates that an important component of the persistence strategy involves active interference with the MHC class I antigen processing pathway during the lytic replication cycle. We have now identified a novel role for the lytic cycle gene, BILF1, which encodes a glycoprotein with the properties of a constitutive signaling G-protein-coupled receptor (GPCR). BILF1 reduced the levels of MHC class I at the cell surface and inhibited CD8+ T cell recognition of endogenous target antigens. The underlying mechanism involves physical association of BILF1 with MHC class I molecules, an increased turnover from the cell surface, and enhanced degradation via lysosomal proteases. The BILF1 protein of the closely related CeHV15 c1-herpesvirus of the Rhesus Old World primate (80% amino acid sequence identity) downregulated surface MHC class I similarly to EBV BILF1. Amongst the human herpesviruses, the GPCR encoded by the ORF74 of the KSHV c2-herpesvirus is most closely related to EBV BILF1 (15% amino acid sequence identity) but did not affect levels of surface MHC class I. An engineered mutant of BILF1 that was unable to activate G protein signaling pathways retained the ability to downregulate MHC class I, indicating that the immune-modulating and GPCR-signaling properties are two distinct functions of BILF1. These findings extend our understanding of the normal biology of an important human pathogen. The discovery of a third EBV lytic cycle gene that cooperates to interfere with MHC class I antigen processing underscores the importance of the need for EBV to be able to evade CD8+ T cell responses during the lytic replication cycle, at a time when such a large number of potential viral targets are expressed

A CD8+ T cell immune evasion protein specific to Epstein-Barr virus and its close relatives in Old World primates

γ1-Herpesviruses such as Epstein-Barr virus (EBV) have a unique ability to amplify virus loads in vivo through latent growth-transforming infection. Whether they, like α- and β-herpesviruses, have been driven to actively evade immune detection of replicative (lytic) infection remains a moot point. We were prompted to readdress this question by recent work (Pudney, V.A., A.M. Leese, A.B. Rickinson, and A.D. Hislop. 2005. J. Exp. Med. 201:349–360; Ressing, M.E., S.E. Keating, D. van Leeuwen, D. Koppers-Lalic, I.Y. Pappworth, E.J.H.J. Wiertz, and M. Rowe. 2005. J. Immunol. 174:6829–6838) showing that, as EBV-infected cells move through the lytic cycle, their susceptibility to EBV-specific CD8+ T cell recognition falls dramatically, concomitant with a reductions in transporter associated with antigen processing (TAP) function and surface human histocompatibility leukocyte antigen (HLA) class I expression. Screening of genes that are unique to EBV and closely related γ1-herpesviruses of Old World primates identified an early EBV lytic cycle gene, BNLF2a, which efficiently blocks antigen-specific CD8+ T cell recognition through HLA-A–, HLA-B–, and HLA-C–restricting alleles when expressed in target cells in vitro. The small (60–amino acid) BNLF2a protein mediated its effects through interacting with the TAP complex and inhibiting both its peptide- and ATP-binding functions. Furthermore, this targeting of the major histocompatibility complex class I pathway appears to be conserved among the BNLF2a homologues of Old World primate γ1-herpesviruses. Thus, even the acquisition of latent cycle genes endowing unique growth-transforming ability has not liberated these agents from evolutionary pressure to evade CD8+ T cell control over virus replicative foci

Stage-Specific Inhibition of MHC Class I Presentation by the Epstein-Barr Virus BNLF2a Protein during Virus Lytic Cycle

gamma-herpesvirus Epstein-Barr virus (EBV) persists for life in infected individuals despite the presence of a strong immune response. During the lytic cycle of EBV many viral proteins are expressed, potentially allowing virally infected cells to be recognized and eliminated by CD8+ T cells. We have recently identified an immune evasion protein encoded by EBV, BNLF2a, which is expressed in early phase lytic replication and inhibits peptide- and ATP-binding functions of the transporter associated with antigen processing. Ectopic expression of BNLF2a causes decreased surface MHC class I expression and inhibits the presentation of indicator antigens to CD8+ T cells. Here we sought to examine the influence of BNLF2a when expressed naturally during EBV lytic replication. We generated a BNLF2a-deleted recombinant EBV (ΔBNLF2a) and compared the ability of ΔBNLF2a and wild-type EBV-transformed B cell lines to be recognized by CD8+ T cell clones specific for EBV-encoded immediate early, early and late lytic antigens. Epitopes derived from immediate early and early expressed proteins were better recognized when presented by ΔBNLF2a transformed cells compared to wild-type virus transformants. However, recognition of late antigens by CD8+ T cells remained equally poor when presented by both wild-type and ΔBNLF2a cell targets. Analysis of BNLF2a and target protein expression kinetics showed that although BNLF2a is expressed during early phase replication, it is expressed at a time when there is an upregulation of immediate early proteins and initiation of early protein synthesis. Interestingly, BNLF2a protein expression was found to be lost by late lytic cycle yet ΔBNLF2a-transformed cells in late stage replication downregulated surface MHC class I to a similar extent as wild-type EBV-transformed cells. These data show that BNLF2a-mediated expression is stage-specific, affecting presentation of immediate early and early proteins, and that other evasion mechanisms operate later in the lytic cycle

The Epstein-Barr Virus Glycoprotein gp150 Forms an Immune-Evasive Glycan Shield at the Surface of Infected Cells

Cell-mediated immunity plays a key role in host control of viral infection. This is exemplified by life-threatening reactivations of e.g. herpesviruses in individuals with impaired T-cell and/or iNKT cell responses. To allow lifelong persistence and virus production in the face of primed immunity, herpesviruses exploit immune evasion strategies. These include a reduction in viral antigen expression during latency and a number of escape mechanisms that target antigen presentation pathways. Given the plethora of foreign antigens expressed in virus-producing cells, herpesviruses are conceivably most vulnerable to elimination by cell-mediated immunity during the replicative phase of infection. Here, we show that a prototypic herpesvirus, Epstein-Barr virus (EBV), encodes a novel, broadly acting immunoevasin, gp150, that is expressed during the late phase of viral replication. In particular, EBV gp150 inhibits antigen presentation by HLA class I, HLA class II, and the non-classical, lipid-presenting CD1d molecules. The mechanism of gp150-mediated T-cell escape does not depend on degradation of the antigen-presenting molecules nor does it require gp150’s cytoplasmic tail. Through its abundant glycosylation, gp150 creates a shield that impedes surface presentation of antigen. This is an unprecedented immune evasion mechanism for herpesviruses. In view of its likely broader target range, gp150 could additionally have an impact beyond escape of T cell activation. Importantly, B cells infected with a gp150-null mutant EBV displayed rescued levels of surface antigen presentation by HLA class I, HLA class II, and CD1d, supporting an important role for iNKT cells next to classical T cells in fighting EBV infection. At the same time, our results indicate that EBV gp150 prolongs the timespan for producing viral offspring at the most vulnerable stage of the viral life cycle

Two novel human cytomegalovirus NK cell evasion functions target MICA for lysosomal degradation

NKG2D plays a major role in controlling immune responses through the regulation of natural killer (NK) cells, αβ and γδ T-cell function. This activating receptor recognizes eight distinct ligands (the MHC Class I polypeptide-related sequences (MIC) A andB, and UL16-binding proteins (ULBP)1–6) induced by cellular stress to promote recognition cells perturbed by malignant transformation or microbial infection. Studies into human cytomegalovirus (HCMV) have aided both the identification and characterization of NKG2D ligands (NKG2DLs). HCMV immediate early (IE) gene up regulates NKGDLs, and we now describe the differential activation of ULBP2 and MICA/B by IE1 and IE2 respectively. Despite activation by IE functions, HCMV effectively suppressed cell surface expression of NKGDLs through both the early and late phases of infection. The immune evasion functions UL16, UL142, and microRNA(miR)-UL112 are known to target NKG2DLs. While infection with a UL16 deletion mutant caused the expected increase in MICB and ULBP2 cell surface expression, deletion of UL142 did not have a similar impact on its target, MICA. We therefore performed a systematic screen of the viral genome to search of addition functions that targeted MICA. US18 and US20 were identified as novel NK cell evasion functions capable of acting independently to promote MICA degradation by lysosomal degradation. The most dramatic effect on MICA expression was achieved when US18 and US20 acted in concert. US18 and US20 are the first members of the US12 gene family to have been assigned a function. The US12 family has 10 members encoded sequentially through US12–US21; a genetic arrangement, which is suggestive of an ‘accordion’ expansion of an ancestral gene in response to a selective pressure. This expansion must have be an ancient event as the whole family is conserved across simian cytomegaloviruses from old world monkeys. The evolutionary benefit bestowed by the combinatorial effect of US18 and US20 on MICA may have contributed to sustaining the US12 gene family

Ubiquitylation in ERAD: Reversing to Go Forward?

Proteins are co-translationally inserted into the endoplasmic reticulum (ER) where they undergo maturation. Homeostasis in the ER requires a highly sensitive and selective means of quality control. This occurs through ER-associated degradation (ERAD).This complex ubiquitin-proteasome–mediated process involves ubiquitin conjugating enzymes (E2) and ubiquitin ligases (E3),lumenal and cytosolic chaperones, and other proteins, including the AAA ATPase p97 (VCP; Cdc48 in yeast). Probing of processes involving proteasomal degradation has generally depended on proteasome inhibitors or knockdown of specific E2s or E3s. In this issue of PLoS Biology, Ernst et al. demonstrate the utility of expressing the catalytic domain of a viral deubiquitylating enzyme to probe the ubiquitin system. Convincing evidence is provided that deubiquitylation is integral to dislocation of ERAD substrates from the ER membrane. The implications of this work for understanding ERAD and the potential of expressing deubiquitylating enzyme domains for studying ubiquitin-mediated processes are discussed