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.

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.

Transforming activity of green monkey SA7(C8) adenovirus in tissue culture.

Simian SA7 (C8) adenovirus can effect transformation of mouse, rat, and hamster cells in tissue culture. The transforming activity of the virus was more pronounced in culture medium containing 0.1 millimolar CaCl(2), a lower concentration than that usually used. The transformed hamster cells were tumorigenic in hamsters.

Activation of viruses in human tumors by 5-iododeoxyuridine and dimethyl sulfoxide.

Dimethyl sulfoxide added to cultures first treated with 5-iododeoxyuridine increased C-type virus production approximately tenfold in a human rhabdomyosarcoma cell line. 5-Iododeoxyuridine followed by dimethyl sulfoxide also activated a similar C-type virus in a metastatic tumor from a bronchial node taken from a 52-year-old male.

Simian tumor virus isolate: demonstration of cytopathic effects in vitro.

Several cell lines that were derived from primates and inoculated with virus originally obtained from a spontaneous mammary carcinoma showed cytopathic effects characterized by multinucleation. These cytopathic effects appeared as early as 24 holurs after inoculation. Multinucleated cells contained virus particles characteristic of the original virus isolate.

Type C virus from cell cultures of chemically induced rat hepatomas.

Type C RNA viruses are present in cell cultures from transplantable and primary hepatomas induced by aromatic amine carcinogens. Virus yield was markedly enhanced by treating the cells with bromodeoxyuridine. Preparations of rat hepatoma-associated virus obtained from cultures treated with this compound were deficient in DNA polymerase activity.

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.

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.

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.

Replication of feline sarcoma virus in cat embryonic kidney cells in the absence of transformation.

The infection of cat embryonic kidney cells (CRC) with Gardner and Snyder-Theilen strains of feline sarcoma viruses (Ga-FeSV and ST-FeSV) do not lead to transformation and focus formation. The focus forming insusceptibility is shown to be stable and nontransmissable property of these cells. However, the CRC cells are permissive for ST-FeSV, and the viral production is 10 to 100 folds lower than in focus forming susceptible cat cells. Produced ST-FeSV(c) by CRC cells induced on cat embryonic cells FE in higher proportion round and fusiform (rf) foci and fusiform(f) foci. The f foci were not previously observed in original stock of ST-FeSV tested directly on FE cells. Attempts to detect some differences in morphology or in growth behavior between ST-FeSV infected and non-infected CRC cell population gave negative results.

On the problem of oncogene of tumour viruses.

The approach to the problem of oncogenesis of tumorigenic viruses is compared and analyzed from the position of the Altshtein-Vogt hypothesis and from that of the general theory of oncogenesis advanced by the present author. In contrast to the hypothesis of Altshtein-Vogt dealing mainly with the problem of oncogene origin, the general theory of oncogenesis not only defines concretely the origin of the oncogene and the essence of its product, but also makes it possible to understand why, when and how integration of the oncogene with the genome of the cell leads to the transformation of the cell into a benign cell and when into a malignant tumour cell. An analysis of the essence of the "oncogene position effect" from this standpoint shows that an integration, similar in its mechanism but differing in polarity, of the genome of other viruses with the cell genome should lead to the formation of a corresponding antiviral stable (life-long) immunity or also to the emergence of pseudoautoimmune disease of the type caused by "slow" viruses.

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.

[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.

[Comparative study of the intensity of production of LPV oncornavirus by T-9 cell line and an infected culture of human diploid cells]. / Sravnitel'noe izuchenie intensivnosti produktsii onkornavirusa LPV kletkami linii T-9 i zarazhennoi kul'tury diploidnykh kletok cheloveka

A culture of transformed cells, T-9 line, producing LPV oncornavirus and a culture of human diploid cells (HDC) infected with a cell-free ultrafiltrate of these transformed cells were examined in the electron microscope. The study was aimed at determination of the optimal time and intensity of LPV oncornavirus production by one cell of transformed and infected cultures. LPV oncornavirus production in T-9 cell culture was found to have a pattern of slowly developing, non-intensive two-wave process, whereas in infected HDC culture the process was progressing actively resulting in an increase of the amount of intracytoplasmic type A particles and extracellular type B and C virions by 10- and 15-fold, respectively, within 48- 96 hours for T-9 culture and 96 hours for infected HDC. Cell transformation was found to be apparently preceeded by the infectious process the intensity of which decreased as fibroblast cell culture was transformed into a culture of epithelioid cells.

[Oncornavirus D--an inducer of a new antigen]. / Onkornavirus D--induktor novogo antigena

Oncornavirus D inoculated into chick fibroblast cultures induced in them synthesis of a new antigen which is not incorporated into the virion structure and is absent in cells not infected with this virus. The oncornavirus D newly induced antigen is identical to that found in long-term continuous human HeLa, HEp-2 and J-96 cells. These facts give grounds to believe that the new antigen present in these cells emerged as a consequence of their infection with oncornavirus D. However it remains unknown whether this antigen plays the role of the transforming agent and whether transformation of normal human cells into malignant cells is associated with it.

[Study of LPV oncornavirus reproduction in mixed cell culture]. / Izuchenie reproduktsii onkornavirusa LPV v smeshannoi kul'ture kletok

Electron microscopy and mathematical analysis were used to determine the intensity of LPV oncornavirus production by a single cell in co-cultivated human diploid cells and cells of the continuous T-9 line. Maximum production of intracytoplasmic particles of A type was observed at 48 hours of cultivation, and extracellular virions at 96 hours. Mature virions of B type were more numerous than mature virions of C type in the mixed culture. On the whole the amolnt of mature virions was greater than that of immature ones, and the amount of intracytoplasmic A type particles was considerably greater than that of extracellular particles. By the total amount of production of all virus-specific structures and the two-wave pattern of virus production this mixed culture passaged at a ratio of HDC to T-9 cells 2000 : 1 resembled a culture of T-9 line. Thus, this study indicates that the infected HDC culture at later intervals of cultivation (96 hours) is a more active "producer" of LPV oncornavirus than the main line of T-9 cells or mixed HDC--T-9 culture.