M. Saveria Campo

E5 protein of human papillomavirus type 16 selectively downregulates surface HLA class I

Papillomaviruses have evolved mechanisms that result in escape from host immune surveillance. The E5 protein is expressed early in papillomavirus infection in the deep layers of the infected epithelium. It is localized to the Golgi apparatus (GA) and endoplasmic reticulum. The E5 protein of bovine papillomavirus (BPV) impairs the synthesis and stability of major histocompatibility (MHC) class I complexes and prevents their transport to the cell surface due to retention in the GA. Here we show that human papillomavirus type 16 (HPV‐16) E5 also causes the retention of MHC (HLA) class I complexes in the GA and impedes their transport to the cell surface, which is rescued by treatment with interferon. Unlike BPV E5, HPV‐16 E5 does not affect the synthesis of HLA class I heavy chains or the expression of the transporter associated with antigen processing TAP. These results show that downregulation of surface MHC class I molecules is common to both BPV and HPV E5 proteins. Moreover, we determined that HPV‐16 E5 downregulates surface expression of HLA‐A and HLA‐B, which present viral peptides to MHC class I‐restricted cytotoxic T lymphocytes (CTLs), but not the natural killer (NK) cell inhibitory ligands HLA‐C and HLA‐E. Selective downregulation of cell surface HLA class I molecules may allow the virus to establish infection by avoiding immune clearance of virus‐infected cells by both CTLs and NK cells.

E5 protein of human papillomavirus 16 downregulates HLA class I and interacts with the heavy chain via its first hydrophobic domain.

Human papillomavirus type 16 E5 protein (HPV‐16 E5) is expressed early in papillomavirus infection and is localised primarily in the cell Golgi apparatus (GA) and endoplasmic reticulum. E5 prevents transport of the major histocompatibility class I (MHC I; HLA class I in humans) to the cell surface and retains the complex in the GA. We report that these effects are due, at least in part, to the interaction between E5 and HLA I heavy chain (HC). We also demonstrate that the down‐regulation of surface HLA I and interaction with HC are mediated by the first hydrophobic domain of E5. Although E5 downregulates classical HLA selectively as it does not downregulate non‐classical HLA, the interaction with the HC of classical HLA I is not specific for a particular haplotype of HLA I. This suggests that E5 can interfere with antigen presentation by most, if not all, classical HLA I haplotypes, with potentially serious consequences as the ability of infected cells to present antigenic peptides to effector T cells would be compromised.

Down-regulation of MHC class I by bovine papillomavirus E5 oncoproteins

The papillomavirus E5 protein is localized in the endoplasmic reticulum (ER) and Golgi apparatus (GA) of the host cell. Transformed bovine fibroblasts expressing bovine papillomavirus (BPV) E5 are highly vacuolated and have a much enlarged, distorted and fragmented GA. Major histocompatibility complex class I (MHC I) is processed and transported to the cell surface through the GA. Given the cellular localization of E5 in the GA and the morphologically abnormal GA, we investigated the expression of MHC I in cells transformed by E5 from BPV-1 and BPV-4. Two cell lines were used: bovine cells that also express E6, E7 and activated ras, and NIH3T3 cells that express only E5. In addition, PalF cells acutely infected with a recombinant retrovirus expressing E5 were also examined. In contrast to non-transformed normal cells, or transformed cells expressing other papillomavirus proteins, cells expressing E5 do not express MHC I on their surface, but retain it intracellularly, independently of the presence of other viral or cellular oncogenes, or of whether the cells are long-term transformants or acutely infected. We conclude that expression of E5 prevents expression of MHC I to the cell surface and causes its retention within the cell. In addition, lower amounts of total MHC I heavy chain and of heavy chain RNA are detected in E5-transformed cells than in control cells. As surface expression of another glycosylated membrane protein, the transferrin receptor, is not affected, it appears that E5 targets MHC I with at least a degree of specificity. In papillomavirus lesions this effect would have important implications for antigen presentation by, and immunosurveillance of, virally infected cells.

Evasion of host immunity directed by papillomavirus-encoded proteins.

The nature of the interaction between papillomaviruses (PV) and their infected host has led to the identification of ways in which the viral oncoproteins can transform the infected host cells into cancer cells. As viral persistence is required for malignancy, and persistence requires avoidance of immune attack by the host, defining the relationship between PV and the immune system is also paramount in understanding tumorigenesis. It has emerged that PV have evolved several ways in which to prevent clearance by the host immune system. The limitation of the PV replication cycle to the epithelium, together with low level expression of the virus proteins and an absence of inflammation, minimises the exposure of virus to immune cells. In addition, more recently it has been shown that, like many other viruses, PV can directly subvert the immune response, including interference with the interferon pathway, modulation of antigen presentation, inhibition of interleukin-18 activity and down-regulation of major histocompatibility class I on infected cells. Collectively these mechanisms explain how PV lesions are able to persist for long periods of time in immunocompetent hosts.

The bovine papillomavirus oncoprotein E5 retains MHC class I molecules in the Golgi apparatus and prevents their transport to the cell surface

During papillomavirus infection, the E5 protein localizes in the cell Golgi apparatus and other endomembrane compartments. Cells transformed by E5 do not express major histocompatibility class I complex (MHC I) on the cell surface, while cells transformed by the other transforming proteins E6 and E7 do. In addition, the total amount of both MHC I protein and mRNA is reduced in E5-transformed cells. Here we show that expression of bovine papillomavirus E5 causes the retention of MHC I in the Golgi apparatus, thus preventing its transport to the cell surface. We ascribe this effect to a failure of acidification of the Golgi apparatus, as similar effects are observed in control cells treated with the ionophore monensin. Treatment of E5-transformed cells with either β- or γ-interferon increases the synthesis of MHC I, showing that inhibition of MHC I expression by E5 is not irreversible. However, even after interferon treatment, MHC I, although increased in quantity, is not transported to the cell surface. E5 therefore affects MHC I at several levels, but prevention of MHC I transport to the cell surface appears to be the dominant effect. Lack of surface MHC I would have profound consequences for presentation of viral peptides to the immune system.

Papillomavirus prophylactic vaccines: established successes, new approaches.

ABSTRACT Vaccines against the human papillomaviruses (HPVs) most frequently associated with cancer of the cervix are now available. These prophylactic vaccines, based on virus-like particles (VLPs), are extremely effective, providing protection from infection in almost 100% of cases. However, the vaccines present some limitations: they are effective primarily against the HPV type present in the vaccine, are expensive to produce, and need a cold chain. Vaccines based on the minor capsid protein L2 have been very successful in animal models and have been shown to provide a good level of protection against different papillomavirus types. The potential of L2-based vaccines to protect against many types of HPVs is discussed.

The detection of Bovine Papillomavirus type 1 DNA in flies

BPVs are double stranded DNA viruses that can infect several species other than the natural host, cattle, including equids. In equids, BPV-1, and, less commonly BPV-2, infection gives rise to fibroblastic tumours of the skin. Whilst a causal relationship between BPV-1/2 and equine sarcoids is now well established, how the disease is transmitted is not known. In this study we show BPV-1 DNA can be detected in flies trapped in the proximity of sarcoid-affected animals. Sequence analysis of the BPV-1 LCR from flies indicates that flies harbour BPV-1 LCR sequence variants II and IV which are commonly detected in equine sarcoids. These data suggest that flies may be able to transmit BPV-1 between equids.

The E6E7 oncoproteins of cutaneous human papillomavirus type 38 interfere with the interferon pathway.

Non-melanoma skin cancer is the most frequent malignancy in Caucasian populations. Evidence suggests the involvement of cutaneous Human Papillomavirus (HPV) of the genus beta (beta) in this disease. The ability of E6 and E7 of mucosal HPV to promote cellular transformation and inhibit immune response-related pathways plays a key role in cervical carcinogenesis. beta HPV-38 E6 and E7 display transforming activities in in vitro and in vivo models, but their impact on immune surveillance is unknown. Here we show that HPV-38 E6 and E7 affect the IFN-induced up-regulation of MHC class I. Expression of the two viral proteins in HaCaT keratinocytes led to a decrease of MHC I levels. This down-regulation is associated with a reduction of expression of MHC I heavy chain, of the peptide chaperone TAP and of the STAT-1 downstream effector IRF-1. The down-regulation of these proteins is ultimately due to the inhibition of STAT-1 expression. Analysis of cells expressing either HPV-38 E6 or E7 suggests that these effects are primarily the result of E6 expression, although a contribution by E7 cannot be excluded. We conclude that HPV-38 encodes oncoproteins that potentially contribute to the evasion of host immune surveillance.

Papillomaviruses: a correlation between immune evasion and oncogenicity?

Abstract There is increasing evidence that the immune system plays a pivotal role in determining the outcome of papillomavirus infection, including the discovery that papillomaviruses, like other viruses, have evolved mechanisms to avoid immune attack directly. Although, in most cases, the immune system is able to overcome the evasion mechanisms and clear infection, continued persistence of the virus can lead to malignancy. Much research has focused on determining the differences between the so-called high risk (most commonly associated with cancer) and low risk (causing benign infections) human papillomavirus types in their ability to cause cancer, particularly the functional effects of the virus oncoproteins on cellular pathways. Here, we suggest that the ability to avoid immune attack is also linked to the transforming potential of papillomaviruses. We discuss whether those papillomaviruses associated with cancer have an enhanced ability to avoid the immune response compared with their benign-disease-causing counterparts.

Synergism between bovine papillomavirus type 4 and the flavonoid quercetin in cell transformation in vitro

Bovine papillomavirus type 4 (BPV-4) morphologically transforms primary bovine cells in vitro only in the presence of an activated ras gene. The transformed cells are capable of anchorage-independent growth, but are not immortal and are incapable of inducing tumors in nude mice, suggesting that other events are needed to convert the cells to the fully transformed phenotype. We show here that treatment of the cells with a single dose of the flavonoid quercetin leads to full oncogenic transformation of cells transfected with BPV-4 and ras. Quercetin is one of the most potent mutagens found in bracken fern, the environmental cofactor in BPV-4-associated carcinogenesis of the upper alimentary canal of cattle. Our results point to quercetin as the probable in vivo cocarcinogen synergizing with BPV-4 in malignant progression.