Rous Virus Research Articles

Similar Structural Models of the Transmembrane Proteins of Ebola and Avian Sarcoma Viruses

Similar Structural Models of the Transmembrane Proteins of Ebola and Avian Sarcoma Viruses

Rescue of infectious virus from nonproducer Rous cells by chick cellular DNA.

The rescue of infectious virus from nonproducer BH RSV(-) cells by chick cellular DNA was attempted in order to investigate the functional state of endogenous and exogenous retroviral genes integrated within the cellular DNA. No infectious virus was rescued by transfection with DNAs of chick helper factor (chf)-negative chick embryo cells (CEC), chf-positive CEC or uninfected CEC producing endogenous Rous associated virus (RAV-0). On the other hand, infectious Rous viruses with the phenotype of RAV-0 and RAV-1 were rescued by transfection with DNAs of CEC which had been infected with RAV-0 and RAV-1. From these results, it seems that exogenous retroviral genes integrated in the cellular DNA are expressed rather easily by transfection while those present endogenously are not.

Changes in rous virus and polyoma virus during their synthesis in cells of unnatural hosts

A line of human embryonic fibroblasts, transformed by Rous sarcoma virus (Schmidt-Ruppin strain) contained and produced Rous virus, as was shown by the complement fixation and immunofluorescent tests, by electron-microscopic investigation, and by the presence of an isotope peak in a sucrose gradient. By its biological properties the synthesized virus differed from the original Schmidt-Ruppin strain for, in particular, the range of cells sensitive to the virus and its protein membrane were changed. Similar data indicating a change in the biological properties of a virus produced in the tissue of an unnatural host also were obtained for polyoma virus, synthesized in human embryonic fibroblasts transformed by it.

Viral adenosine triphosphatase

The catalytic and immunological properties of an adenosine triphosphatase from different types of virus have been studied. The avian myeloblastosis virus has been found to be specialized in holding this enzyme in a highly active state as compared to other virus with respect to their host cell enzyme. Catalytically myeloblastosis virus and Rous virus ATPase behave alike, while that of the Reo virus is significantly different.

Cytoplasmic vacuoles of Rous virus transformed cells are organelles involved in cation uptake.

Cytoplasmic vacuoles induced during transformation of cells by Bryan strain Rous sarcoma virus (RSV-BH) have been studied using the cationic dye, neutral red(NR). Both the rate of uptake and the accumulation of NR are greater in RSV-BH transformed cells than non-transformed cells however, uptake was greater in vacuolated than in non-vacuolated cells, whether or not they were transformed. The NR was incorporated into pre-existing vacuoles in the absence of cytoplasmic staining, suggesting the existence of direct channels from the cell surface to the vacuoles. Other low mol. wt. cationic dyes could also be incorporated into vacuoles, although those with branched structures or cationic weights greater than 330 were excluded. No anionic dyes were incorporated. Infection of cells with a virus mutant, RSV-BH-Ta, induces temperature-dependent vacuolization. After a shift to the vacuole-permissive temperature, vacuoles developed at different rates and with morphological variations with different cations. Vacuoles which had formed in the presence of several cations, (K+, Rb+, tris+, choline+) failed to disappear when cells were incubated at a temperature sufficient to revert vacuoles formed in Na+-containing medium. No short-term effects of Cl-replacements (Br-, I-, or SO2-4) on vacuolization or reversal were observed. The results suggest that these vacuoles are organelles involved in cation uptake. A possible function for these organelles in RSV-BH induced malignancy is discussed.

Incomplete viral genome in a non‐virogenic mouse tumour cell line (RVP3) transformed by prague strain of avian sarcoma virus

Two cell lines, RVP3 and RVA4, derived originally from mouse tumors induced by the Prague and Schmidt-Ruppin strain of RSV, respectively, were studied. tall attempts failed to induce infectious virus production in RVP3 cells by fusion with chicken fibroblasts even if the cells were infected with avian leukosis viruses. Also, attempts to rescue the viral genome by transfection were unsuccessful. RVP3 cells harboured 31-45% of the viral genome sequences, as was shown by molecular hybridization, and therefore they were designated cryptovirogenic. The tumour cell line RVA4 did not contain any detectable viral sequences. The significance of the detection of the incomplete Rous virus genome sequences in mammalian cells is discussed.

Presence of Chicken Cell Surface Antigen on Rous Virus Activated in Heterokaryons of Transformed Non-permissive Hamster Cells and Chicken Cells

Incubation with antiserum to chick embryo (CE) cells, in the presence of complement, inactivates Rous sarcoma virus (RSV) produced by heterokaryons formed by non-permissive RSV-transformed hamster cells and CE cells as well as RSV produced by heterokaryons is observed following incubation with antiserum to the transformed hamster cells, plus complement. Hence, the envelope of RSV activated in heterokaryons, as that of RSV produced by CE cells, must contain a surface antigen of the CE cell, and the virus must mature only, or preferentially, at chicken-specific sites of the heterokaryon surface.

Ultrastructural study of heterokaryons from rous rat sarcoma cells and normal chickens cells.

Rous virus transformed rat cells containing the Rous virus genome but lacking the ability to produce Rous virus particles have been fused with chicken erythrocytes, chicken lymphocytes or chicken fibroblasts using Sendai virus. Heterokaryotic cells with both rat sarcoma cell nuclei and chicken cell nuclei have been studied in the electron microscope. Shortly after the fusion, a series of changes in the chicken nuclei are demonstrable, which as seen in the electron microscope imply a “reactivation” and production of intranuclear RNA‐containing structures as well as a development of nucleoli. In parallel, the chicken nucleus increases in size and the chromatin is transformed from condensed aggregates into a finely dispersed material. In spite of all evidence of a protein synthesis in the heterokaryotic Rous sarcoma cell directed from the chicken nucleus there is no formation of virus particles in cells with Rous rat sarcoma cells and chicken erythrocytes or chicken lymphocytes. In cultures with Rous rat sarcoma cell—chicken fibroblast heterokaryons a regular synthesis was seen. It seems apparent that the chicken fibroblast contains some factor, whether genetically coded or not, that is required if a production of complete Rous virus particles is to be achieved.

Selective incorporation of host cell methionyl-transfer RNA by RNA tumor viruses

The low molecular weight RNA in three strains of avian tumor viruses, namely, Rous associated virus-1, Bryan Rous sarcoma virus grown in the presence of Rous associated virus-1, and Schmidt-Ruppin Rous sarcoma virus, accepted a large number of amino acids. In all three strains of Rous viruses, transfer RNA for methionine was incorporated in relatively large amounts. An examination of the reversed-phase-5 chromatographic profiles of methionyl-tRNA from the viruses and host cells demonstrated that one isoaccepting species of methionyl-tRNA was preferentially incorporated by all three viruses from the host cells. Transformylation and ribosome binding experiments indicated that this species was the internal methionyl-tRNA rather than the initiating species of formylatable methionyl-tRNA.

Further studies on the ability of regressor sera to block cell-mediated destruction of Rous sarcomas.

AbstractPrevious studies have shown that most sera from Japanese quails with growing sarcomas induced by the Schmidt‐Ruppin strain of the Rous virus (SR‐RSV) can block cell‐mediated destruction of Rous sarcoma cells in vitro, and that sera from some quails whose Rous sarcomas have spontaneously regressed (regressors) also have that ability. Six regressor sera, previously shown to be blocking when added to the mixture of target and effector (=regressor spleen) cells in vitro, were tested for blocking effect at either the target or the effector cell level. Three sera could block only when incubated with the target cells, while three other sera blocked at both the target cell and the effector cell level. Two of three regressor sera which had previously been shown to be unblocking (abrogating the blocking activity of progressor sera when mixed with these in vitro) increased cell‐mediated cytotoxicity at a low dilution (potentiation) but had a slight blocking activity at dilution 1:128–1:512, which disappeared upon further dilution. The blocking effect seen at these dilutions appeared to occur at the target cell level and was not detectable after incubation of the effector cells with serum. After incubation of the target cells for 24 h with one of these two regressor sera diluted 1:256, a blocking activity detectable at the effector cell level appeared in the culture medium. The blocking activity of serum from bursectomized progressors (previously reported) increased, after regressor serum diluted 1:16 had been added to it; at that dilution, the regressor serum had no blocking activity by itself. More concentrated regressor serum, on the other hand, was unblocking, i.e. it decreased the blocking activity of serum from bursectomized progressors. The data are compatible with the hypothesis that the blocking factors operating in this system are complexes between antigens released from the tumor and antibodies formed by the host (and, occasionally, free antigen).

Ribonucleases of Rous sarcoma virus

Purified Rous sarcoma virus and avian leukosis virus preparations contain ribonucleases which hydrolyze polyuridylic acid (Poly U), polyadenylic acid (Poly A) and the double-stranded hybrid of poly U and poly A (Poly A:Poly U). The ribonucleases become soluble when the virus is dissociated with a neutral detergent (Brij 35), mercaptoethanol, and urea. Carboxymethyl cellulose chromatography partially separates the ribonucleases into two activity peaks; one for Poly U and the other for Poly A or Poly A:Poly U. The chromatographed fractions are further purified by gel filtration yielding approximate molecular weights of 13,000 for the enzymes. The partially purified enzyme preparations also digest the viral RNA in disrupted virions, but not the viral DNA in the disrupted virions synthesized by reverse transcriptase of the virus. The intact virus in solution shows a very high ribonuclease activity toward Poly U, but not toward Poly A or Poly A:Poly U. The viral RNA in the intact virus is stable during a prolonged incubation in spite of the fact that the virus contains the ribonucleases capable of digesting the RNA. Moreover, the viral RNA in the disrupted virions is also stable when incubated alone, although it becomes digested when the viral enzyme preparation is added. These observations suggest that the ribonuclease which prefers Poly U is located on the surface, and that the other ribonucleases which prefer Poly A and Poly A:Poly U reside between inner and outer membranes of the virion. The origin of the ribonucleases in Rous virus is not clear. From chick embryo fibroblasts, ribonucleases for Poly U, Poly A and Poly A:Poly U can be isolated with carboxymethyl cellulose chromatography. The ribonuclease activity for the double-stranded RNA is low in these preparations. After transformation by Rous virus, the chick cells demonstrate all these ribonuclease activities with a considerable increase in the activity for Poly A:Poly U. The ribonuclease activities of purified Rous virus resemble those of the transformed cells.

Evidence for 30-40S RNA as precursor of the 60-70S RNA of Rous sarcoma virus.

Rous sarcoma virus harvested from cells at intervals of 3 min has the same density, sedimentation coefficient, and DNA polymerase as virus harvested at hourly intervals. The RNA of the Prague strain-C consists of a minor class of 60-70S RNA, a major class 30-40S RNA, and a 4-12S class of RNA present at variable concentration. The RNA of the Schmidt-Ruppin strain-A contains more 60-70S than 30-40S RNA. Upon incubation of virus harvested at 3-min intervals at 40 degrees in cell growth medium or Tris-saline, most of the 30-40S RNA is converted to 60-70S RNA. The electrophoretic mobility of the 30-40S RNA of the Rous virus harvested at 3-min intervals is lower than that of the 30-40S subunits of completely dissociated 60-70S RNA; after heating, their mobilities are identical. Heating also releases some small RNAs from 30-40S RNA of virus harvested at 3-min intervals, but five times more 4S RNA is released if the 30-40S RNA is allowed to convert to 60-70S in the virus. The template activity for Rous virus DNA polymerase of the 30-40S RNA of Rous virus harvested at 3-min intervals is about five times lower than that of 60-70S RNA. It is suggested that association of 30-40S RNAs with some RNAs of the 4-12S class may take place simultaneously with their conversion to 60-70S RNA.

Rous sarcomas induced by heterotransplantation of virogenic rat tumour cells in mice.

AbstractThe virogenic sarcoma RSL was induced in inbred Lewis rats by the Schmidt‐Ruppin variant of Rous virus (SR‐RSV).Heterotransplantation of RSL cells to newborn C57BL/10Sn mice gave rise to mouse sarcomas in approximately 10% of the recipients. The properties of one of these mouse sarcomas, designated as RSM, were compared with those of the original rat tumour line RSL.It was found that (1) RSL cells did not contain detectable amounts of infectious chicken, rat or mouse sarcoma virus. However, in contact with permissive chicken fibroblasts the RSL cells produced malignant conversion of avian cells accompanied by the formation of infectious RSV. RSL cells carried a tumour‐associated transplantation antigen (TATA) of Rous sarcomas, a group‐specific (gs) antigen of avian leukoviruses and a membrane antigen (MA), characteristic of Rous sarcomas, detectable by indirect immunofluorescence and cross‐reacting with MA of mouse Rous sarcomas. The MA characteristic of tumours induced by mouse leukoviruses was not detected in the membrane of RSL cells.(2) RSM cells did not contain detectable amounts of oncogenic avian, mouse or rat viruses in an infectious form. Neither contact of these cells with permissive chicken c/o fibroblasts nor heterotransplantation of RSM tumours to chickens led to malignant conversion of avian cells. Gs antigen of avian leukoviruses was not detectable in RSM cells. However, TATA and MA characteristic of Rous sarcomas were present. Neither MA nor TATA induced by mouse leukoviruses was detected in RSM cells.The results obtained indicate that the non‐infectious RSV genome present in rat virogenic cells is able to induce tumours in a further mammalian species.

Immunological study of tumors induced by highly oncogenic forms of rous virus in adult mice

Immunological study of tumors induced by highly oncogenic forms of rous virus in adult mice

A strain of hamster embryonic fibroblasts transformed by rous virus in vitro

A strain of hamster embryonic fibroblasts transformed by rous virus in vitro

Infective properties of Rous virus RNA and of total RNA from tumors induced by this virus

Infective properties of Rous virus RNA and of total RNA from tumors induced by this virus

Transformation of human cells by polyoma and Rous sarcoma viruses mediated by inactivated Sendai virus.

AbstractTransformation of embryonic human fibroblasts was obtained by treatment with inactivated Sendai virus and polyoma virus. Transformed cells differed morphologically from normal cells and formed a stable cell line (P‐2). A stable transformed cell line (23) was also formed after absorption of Rous sarcoma virus (Schmidt‐Ruppin strain) and inactivated Sendai virus on the embryonic human fibroblasts.Cell line P‐2 and cell line 23 contained a human chromosome set, although some karyological abnormalities were apparent.The cell lines P‐2 and 23 showed some characteristics of malignant cells growing in vitro (ability to multiply when suspended in semi‐solid medium and to form tumours in the cheek pouches and brains of golden hamsters). The potent transforming factor (s) was/were synthesized by these cells. The transforming factors had some properties identical to those of polyoma or Rous viruses and caused rapid transformation of embryonic mouse and human cells growing in vitro.Foci of epithelioid cells appeared in human embryonic fibroblasts after the absorption of human adenovirus type 12 and inactivated Sendai virus.

Antigens of Rous virus particles in virus-free tumors induced in adult rats

Antigens of Rous virus particles in virus-free tumors induced in adult rats

Activation of Rous virus in the transplanted golden hamster tumour with the aid of artificial heterokaryon formation.

AbstractA transplanted hamster tumour induced by the Carr‐Zilber strain of Rous sarcoma virus was studied during ten consecutive passages. Mature virus, virus connected with the cells, viral antigen and antigen active in the complement fixation test were determined. The ability of Rous virus to be activated in tumour cells was also determined with the aid of heterokaryon formation with chick embryo fibroblasts.It was shown that the tumour retained in all passages the capacity for activation of Rous virus despite the disappearance of virus and viral antigen. The minimum number of cells containing one activated virus unit was increased during the tumour passages. X‐irradiation of the tumour cells did not activate the Rous virus.The method of heterokaryon formation was also successfully applied for activation of Rous virus from a cell line that had originated in a hamster Rous tumour and had been cultivated in vitro for more than two years.

The stimulating effect of complete Freund's adjuvant on tumour induction by polyoma virus in mice and by Rous sarcoma virus in rats.

AbstractComplete Freund's adjuvant was shown to greatly stimulate viral oncogenesis both in the case of polyoma virus in mice and in the case of Rous sarcoma virus in rats. Tumours appeared in 74% of CC57BR/Mv mice infected when newborn with polyoma virus — adjuvant emulsion, whereas control mice injected with virus alone failed to develop tumours. Rous virus (Carr‐Zilber strain) and adjuvant when administered together as emulsions to Wistar rats (newborn or sucklings as well as 2.5‐month‐old animals) resulted in a 100% tumour incidence. In control animals treated with the virus alone when 0–4 days old, tumour incidence was no more than 30–40%; no tumours appeared in control animals which received virus alone when they were more than 2 weeks old. A significant stimulating effect of Freund's adjuvant was also revealed when the virus was inoculated into 4‐ to 15‐day‐old rats which received adjuvants as neonates. Thirty per cent of rats given virus — adjuvant emulsion produced measurable quantities of complement‐fixing antibodies to soluble antigens of Rous sarcoma, while in animals given virus alone such antibodies were not demonstrable. It is suggested that the stimulating action of complete Freund's adjuvant on polyoma and Rous oncogenesis is based upon a mechanism similar to the classical “enhancement phenomenon”.