Expression patterns of endogenous avian retrovirus ALVE1 and its response to infection with exogenous avian tumour viruses.

Endogenous retroviruses (ERVs) are genomic elements that are present in a wide range of vertebrates and have been implicated in a variety of human diseases, including cancer. However, the characteristic expression patterns of ERVs, particularly in virus-induced tumours, is not fully clear. DNA methylation was analysed by bisulfite pyrosequencing, and gene expression was analysed by RT-qPCR. In this study, we first found that the endogenous avian retrovirus ALVE1 was highly expressed in some chicken tissues (including the heart, bursa, thymus, and spleen) at 2 days of age, but its expression was markedly decreased at 35 days of age. In contrast, the CpG methylation level of ALVE1 was significantly lower in heart and bursa at 2 days than at 35 days of age. Moreover, we found that the expression of ALVE1 was significantly inhibited in chicken embryo fibroblast cells (CEFs) and MSB1 cells infected with avian leukosis virus subgroup J (ALVJ) and reticuloendotheliosis virus (REV) at the early stages of infection. In contrast, the expression of the ALVE1 env gene was significantly induced in CEFs and MSB1 cells infected with Marek's disease virus (MDV). However, the methylation and expression levels of the ALVE1 long terminal repeat (LTR) did not show obvious alterations in response to viral infection. The present study revealed the expression patterns of ALVE1 in a variety of chicken organs and tissues and in chicken cells in response to avian tumour virus infection. These findings may be of significance for understanding the role and function of ERVs that are present in the host genome.

Construction of a molecular clone of ovine enzootic nasal tumor virus.

BACKGROUND: Enzootic nasal tumor virus (ENTV-1) is an ovine betaretrovirus that has been linked to enzootic nasal adenocarcinoma (ENA), a contagious tumor of the ethmoid turbinates of sheep. Transmission experiments performed using virus isolated from cell free nasal tumor homogenates suggest that ENTV-1 is the causative agent of ENA; however, this etiological relationship has not been conclusively proven due to the fact that the virus cannot be propagated in vitro nor is there an infectious molecular clone of the virus. METHODS: Here we report construction of a molecular clone of ENTV-1 and demonstrate that transfection of this molecular clone into HEK 293T cells produces mature virus particles. RESULTS: Analysis of recombinant virus particles derived from the initial molecular clone revealed a defect in the proteolytic processing of Gag; however, this defect could be corrected by co-expression of the Gag-Pro-Pol polyprotein from the highly related Jaagsiekte sheep retrovirus (JSRV) suggesting that the polyprotein cleavage sites in the ENTV-1 molecular clone were functional. Mutagenesis of the molecular clone to correct amino acid variants identified within the pro gene did not restore proteolytic processing; whereas deletion of one proline residue from a polyproline tract located in variable region 1 (VR1) of the matrix resulted in production of CA protein of the mature (cleaved) size strongly suggesting that normal virion morphogenesis and polyprotein cleavage took place. Finally, electron microscopy revealed the presence of spherical virus particles with an eccentric capsid and an average diameter of about 100 nm. CONCLUSION: In summary, we have constructed the first molecular clone of ENTV-1 from which mature virus particles can be produced. Future experiments using virus produced from this molecular clone can now be conducted to fulfill Koch's postulates and demonstrate that ENTV-1 is necessary and sufficient to induce ENA in sheep.

Immunodeficiency-associated viral oncogenesis.

Several viruses with different replication mechanisms contribute to oncogenesis by both direct and indirect mechanisms in immunosuppressed subjects after solid organ transplantation, after allogeneic stem cell transplantation, or with human immunodeficiency virus (HIV) infection. Epstein-Barr virus (EBV), human papillomavirus (HPV), Kaposi sarcoma herpesvirus (KSHV), human T-cell lymphotropic virus type 1 (HTLV-1) and Merkel cell polyoma virus (MCV) are the main viruses associated with the development of cancer in immunosuppressed patients. Besides being a main cause of immunodeficiency, HIV1 has a direct pro-oncogenic effect. In this review, we provide an update on the association between the condition of acquired immunodeficiency and cancer risk, specifically addressing the contributions to oncogenesis of HPV, MCV, KSHV, HTLV-1, and EBV.

The modulation of apoptosis by oncogenic viruses.

Transforming viruses can change a normal cell into a cancer cell during their normal life cycle. Persistent infections with these viruses have been recognized to cause some types of cancer. These viruses have been implicated in the modulation of various biological processes, such as proliferation, differentiation and apoptosis. The study of infections caused by oncogenic viruses had helped in our understanding of several mechanisms that regulate cell growth, as well as the molecular alterations leading to cancer. Therefore, transforming viruses provide models of study that have enabled the advances in cancer research. Viruses with transforming abilities, include different members of the Human Papillomavirus (HPV) family, Hepatitis C virus (HCV), Human T-cell Leukemia virus (HTLV-1), Epstein Barr virus (EBV) and Kaposi's Sarcoma Herpesvirus (KSHV).Apoptosis, or programmed cell death, is a tightly regulated process that plays an important role in development and homeostasis. Additionally, it functions as an antiviral defense mechanism. The deregulation of apoptosis has been implicated in the etiology of diverse diseases, including cancer. Oncogenic viruses employ different mechanisms to inhibit the apoptotic process, allowing the propagation of infected and damaged cells. During this process, some viral proteins are able to evade the immune system, while others can directly interact with the caspases involved in apoptotic signaling. In some instances, viral proteins can also promote apoptosis, which may be necessary for an accurate regulation of the initial stages of infection.

Postulation of and evidence for provirus existence in RSV-transformed cells and for an oncogenic activity associated with only part of the RSV genome.

This article gives a historical insight into the establishment of suitable models allowing the postulation that chicken Rous sarcoma virus (RSV) becomes integrated in different cells as a provirus. This is documented by the correspondence between two laboratories involved in these investigations. Special attention is paid to RSV-transformed mammalian cells, their virogenic nature, virus rescue by cell fusion, and finally their use for the oncogene v-src characterization. Two sets of experiments are mentioned, which provided an early indication of a transforming gene present in RSV.

Reflections on Epstein-Barr virus: some recently resolved old uncertainties.

The change over recent decades in perceptions of the role of viruses in human cancer-causation is illustrated by the reception given to the discovery of Epstein-Barr virus (EBV) in 1964 compared to that of Kaposi's sarcoma herpesvirus (KSHV or HHV-8) in 1994. Very new data on EBV-like agents in New World monkeys is considered in relation to the antiquity of the association of proto-EBV with early anthropoids. Although the finding that individuals without B lymphocytes do not seem to be infected with EBV appears to have resolved the controversy regarding the permissive cell type producing infectious virus in the oropharynx, the presence of EBV in certain squamous and other epithelial cells raises continuing problems which are discussed. Among many recent successes of molecular biology applied to EBV, new information from such investigations on the genetic defect in X-linked lymphoproliferative syndrome now explains the cause of the disastrous pathological changes underlying the disease.Finally, current progress with vaccines against EBV is reviewed.

cAMP-independent activation of the adenovirus type 12 E2 promoter correlates with the recruitment of CREB-1/ATF-1, E1A(12S), and CBP to the E2-CRE.

Expression of the transcription unit early region 2 (E2) is of crucial importance for adenoviruses because this region encodes proteins essential for viral replication. Here, we demonstrate that the E1A(12S) protein of the oncogenic adenovirus serotype 12 activates the E2 promoter in dependence of the N terminus and the conserved region 1. Activation is mediated through a cAMP-response element that is bound by CREB-1 and ATF-1. Moreover, the Ad12 E2 promoter is inducible by protein kinase A and repressed by either a dominant-negative cAMP-response element-binding protein (CREB) mutant or the highly specific protein kinase A inhibitor protein underscoring the participation of CREB-1/ATF-1 in promoter activation. E1A(12S) binds to CREB-1 and ATF-1 in dependence of the N terminus and CR1 and is recruited to the E2 cAMP-response element through both cellular transcription factors. Most interestingly, point mutations revealed that E1A(12S) domains essential for binding to CREB-1/ATF-1 and for activation of the Ad12 E2 promoter are also essential for binding to the CREB-binding protein. Due to these data and results obtained in DNA-dependent protein-protein interaction assays, we propose a model in which the cAMP-independent activation of the Ad12 E2 promoter is mediated through a ternary complex consisting of CREB-1/ATF-1, E1A(12S), and CREB-binding protein, which assembles on the E2 cAMP-response element.

[Neoplastic disorders and organ transplantation]. / Affections néoplasiques et transplantation d'organe.

INTRODUCTION: Organ transplantation is associated with an increased risk of neoplasia, which seems to be caused by the total effect of immunosuppression, i.e., the combination of factors involved, rather than by the use of a specific class of immunosuppressants. The presence and proliferation of viral oncogenes is frequently observed during this immunosuppressive state. The neoplasia in immunosuppressed patients therefore has particular histological, clinical, evolutive, and therapeutical characteristics. CURRENT KNOWLEDGE AND KEY POINTS: The oncogenic mechanisms in immunosuppressed patients have been progressively clarified. A viral infection is associated with each type of neoplasia: thus, B lymphoma are generally associated with Epstein-Barr viral infection. Skin and uterine cervical carcinomas frequently appear after viral dysplasia due to papillomavirus. The significant increase in the incidence of Kaposi sarcoma shows the role of the immune system in the control of the infection by the human herpes virus 8, which has been recently discovered. Liver cancer is associated with a history of hepatitis B or C chronic infection. FUTURE PROSPECTS AND PROJECTS: Post-transplantation neoplasia constitutes a major problem in patient follow-up, as the number of transplant patients has increased and their survival rate has improved. In addition, there is an increasingly powerful new generation of immunosuppressive drugs. A precise knowledge of the immune system's control mechanisms regarding neoplasic cells and viral infection is an important step in the prevention and efficient treatment of these forms of cancer. Further research into the relationship between the immune system and viral oncogenesis should therefore be considered a major aim.

How viral oncogenes make the cell cycle.

Oncogenic viruses provide their host cells with additional growth stimuli, thereby extending their proliferative capacity. This implies that viral oncogenes can override growth-suppressive signals, which control cell-cycle progression in untransformed cells. Viral oncoproteins deregulate cell-cycle control by interfering with receptor-mediated signal transduction pathways and the function of nuclear cell-cycle regulatory proteins. As a consequence of these regulatory interactions, many viral oncogenes induce the expression of cellular genes required for cell-cycle progression, including genes encoding G1 cyclins. Apparently, different oncogenic viruses target different subsets of these cell-cycle regulatory pathways to transform cells.

The effect of a control program for enzootic bovine leukosis. Changes in herd prevalence in Denmark, 1969-1978.

As part of the Danish control program for enzootic bovine leukosis, three hematologic screenings of all cattle herds in Denmark were performed in the period 1969-1978. Herds with multiple cases of persistent lymphocytosis and/or leukotic tumors were classified as "leukosis herds." During nationwide blood testing, 369 leukosis herds were discovered, and an additional 77 herds were found due to follow-up of tumor cases or special testing in connection with movements of cattle in 1969-1978. In the present study, the prevalence of leukosis herds in the three screening rounds is related to geographic area and herd size. By means of a statistical log-linear model, the authors show that prevalence increased proportionally with herd size. The observed number of leukosis herds per 10,000 herds tested declined during the screening program from 105 to 38 in east Denmark and from 12 to 4 in west Denmark.

Production of human influenza virus in a stabile line of guinea pig tongue cells expressing endogenous oncovirus: an electron microscopic study.

Guinea pig tongue (GPT) cells represent a highly sensitive host system for influenza A/WSN (H1N1) infection as evidenced by numerous ultrastructural changes, considerable production of NS1 protein and widespread budding of viral particles at the cytoplasmic membrane. Vesicles of smooth endoplasmic reticulum and of the Golgi complex were transported to the apical area of cell membrane, where the budding of virions took place. Numerous microtubules were directed vertically to these portions of plasma membrane. In contrast, maturation of the endogenous oncovirus particles occurred at the lateral cytoplasmic membrane. Beneath the area of oncovirus maturation and release, a network was seen of microfilaments oriented towards the plasma membrane. The cytoplasm of GPT cells contained numerous nonstructural protein inclusions, which evidently accumulated at the periphery of nucleoli and were seen to reach the cytoplasm crossing the pores of nuclear membrane.

Persistence of one species of oncovirus in mixed infection after antiserum treatment or transspecies rescue.

Attempts were made to change the envelope of murine sarcoma virus (MSV) from B-MuX (a xenotropic murine leukaemia virus isolate) to feline leukaemia virus (FeLV) by infecting cat cells with MSV(B-MuX), adding excess FeLV as helper and using mouse serum oncovirus-inactivating factor or anti-B-MuX serum. In several attempts, virus from single foci of MSV initially contained only MSV(FeLV) but on passage each showed a minimal persistence of B-MuX. To eliminate the B-MuX component, MSV(B-MuX) was subjected to two consecutive transspecies rescues. The first was performed by co-cultivation of MSV(B-MuX)-producing quail cells with mouse 3T3FL cells, which are completely non-permissive for B-MuX, and pure ecotropic Friend Eveline strain of murine leukaemia virus (F-MuLV); this resulted in apparently pure MSV(F-MuLV). Second, these MSV(F-MuLV)-infected 3T3FL cells were co-cultivated with FeLV and cat cells, which are completely non-permissive for F-MuLV; this resulted in the generation of MSV(FeLV). Passage of this apparently pure FeLV pseudotype in cells permissive only for the replication of B-MuX surprisingly revealed residual murine xenotropic virus. It appears that pressure for survival resulted in genomic masking of B-MuX by heterologous virus envelopes. This phenomenon, which also occurs extensively in nature, implies that if absolute oncovirus genetic purity is required, even extensive attempts at purification may be inadequate.

[Production of noninfectious virions of the Rous sarcoma virus by virogenic hamster cells]. / Produktsiia neinfektsionnykh virionov virusa sarkomy Rausa virogennymi khomiachkovymi kletkami.

Noninfectious virions morphologically identical with avian type C virus virions were produced in Rous sarcoma virus-transformed virogenic hamster cells. The population of the virions contained the major internal protein of avian oncornavirus. It is assumed that production of defective RSV virions occurred in the cells. The major internal protein of avian oncornaviruses was found to be incorporated into the virions containing in their membrane interspecies antigens of hamster oncornavirus produced spontaneously in the system under study. Thus, phenotypic mixing of avian and animal type C viruses in mammalian cells has first been observed.

A marsupial oncovirus?

A virus-like particle was observed in two continuous cell lines derived from the marsupial Sminthopsis crassicaudata (Fat-tailed Dunnart). The development of the particle was similar to the development of D-type oncoviruses. Initially, a crescentof nucleoid material was observed near the nucleus in the region of the Golgi apparatus. This crescent developed into a doughnut-shaped-A-type particle which migrated through the cytoplasm towards the cell membrane where it budded either into a smooth membrane cytoplasmic vacuole or from the cell membrane. Only enveloped A-type particles were observed; no mature B-type, C-type or D-type particles were detected.

Phenotypic mixing between two primate oncoviruses.

Phenotypic mixing between the primate oncoviruses HL23V and BEV has been demonstrated to occur in doubly-infected bat lung (Tb) cells with the production of HL23V(BEV) pseudotype virus. The presence of the HL23V(BEV) pseudotype permitted the host range for replication of HL23V to be extended to murine cells previously 'resistant' to HL23V replication due to a block at the level of virus penetration. Expression of BEV genetic information was observed in doubly-infected rat cells and also in mouse and rat MSV-transformed non-producer cell lines co-cultivated with BEV-producing Tb cells. No evidence for genetic recombination between these viruses could be demonstrated.