[Antimutagenic properties of substances containing beta-carotene]. / Antimutagennye svoistva preparatov, soderzhashchikh beta-karotin.

Two carotene-containing drugs (natural and artificial) were shown to decrease the rate of chromosomal aberrations, induced by cyclophosphane, in bone marrow cells of mice, daily ration of which contained 10-20 mg/kg of beta-carotene within 1-3 weeks. The rate of chromosomal aberrations induced was decreased 1.5-2-fold in various experiments. Artificial beta-carotene exhibited aftereffect within at least 3 days, while its protective effect was decreased about by 20% within 7 days after administration.

[Relation between DNA replication and neutral Mn 2+-dependent DNAse activity in chromatin]. / Zavisimost' replikatsii DNK ot aktivnosti neitral'noi Mn 2+-zavisimoi DNKazy khromatina.

In cultures of primary murine fibroblasts the 10% serum stimulates the replicative synthesis of DNA inhibited by aphidicolin and araC (cytosine arabinoside). Using direct immunofluorescence analysis, it was shown that antibodies penetrate inside the cells and after 4 hours are pooled in the nuclei, where they remain for another 20 hours. The substitution of antibodies against chromatin DNAase by bovine serum albumin of normal serum gamma-globulins does not interfere with the DNA synthesis induction.

[Ability of virus SV40 T-antigen to simulate the effect of specific T lymphocyte growth factor--interleukin-2]. / Sposobnost' T-antigena virusa SV 40 zameniat' deistvie spetsificheskogo rostovogo faktora T-limfotsitov--interleikina-2.

The entering of T-lymphocytes into the DNA-synthesizing phase was marked by three consecutive signals, i.e., antigenic influence, interleukin-2, a specific T-lymphocyte cell growth factor, and non-specific serum growth-promoting factors, in the first place, transferrin. This system was used for the study of effects of virus SV40 T-antigen on cell mitotic cycle. Purified T-antigen was injected consecutively into T-lymphocytes, using erythrocyte ghost vesicles instead of one of control signals. It was shown that T-antigen cannot simulate the antigenic response but simulates the effect of interleukin-2, a specific growth-promoting factor. However, both normally proliferating T-lymphocytes and T-antigen-induced lymphocytes showed an absolute requirement for transferrin and, apparently, for other nonspecific growth-promoting factors. It was assumed that the polymorphism of tumours induced by papovaviruses is determined by the ability of their "early" proteins to imitate the effects of their specific growth-promoting factors on the cells.

[Induction of chromosome aberrations by the T antigen of the SV40 virus introduced into the cells using liposomes]. / Induktsiia khromosomnykh aberratsii T-antigenom virusa SV40, vvedennogo v kletki pri pomoshchi liposom.

A purified SV40 T antigen introduced into hamster cells by means of liposomes accumulated in the nuclei within 10 h and persisted there further as long as 10 to 12 h. Within the first day after cell treatment with T antigen numerous chromosome aberrations including breaks, translocations and gaps were observed in the cells. The number of aberrations slightly reduced by the 2nd day followed by restoration of the normal cell karyotype by the 5th day. Removal of T antigen incorporated into liposomes by a specific immunosorbent or heat inactivation of T antigen abolished the clastogenic effect. It is suggested that induction of chromosome aberrations might activate sell protooncogenes and thus serve a genetic basis of tumor progression.

[Possible role of the T antigen in inducing chromosome aberrations in SV40 virus-transformed cells]. / Vozmozhnaia rol' T-antigena v induktsii khromosomnykh aberratsii v kletkakh, transformirovannykh virusom SV40.

A possible role of the simian virus 40 T antigen in chromosome damages in transformed cells was examined. Two lines of Golden hamster embryonal fibroblasts, transformed by SV40 tsA30 and ts239 mutants (He30 and He239, respectively), were incubated at nonpermissive (40.5-41 degrees C) or permissive (33 degrees C) temperatures. Chromosome aberrations were registered in either subline after 3, 6, 9 and 12 weeks of cultivation under the above conditions. In the both cell lines kept at 33 degrees the frequency of aberrant metaphases and the number of chromosome breaks per cell increased drastically by week 3 of cultivation, and such a state was preserved up to week 12. The frequency of aberrant metaphases in cells cultivated at 41 degrees was maintained at the constant level (He239) or at slightly higher than that in the original culture (He30). The sublines He239, originally incubated at 33 or 40.5 degrees, were then shifted to 40.5 and 33 degrees, respectively. As a result the number of chromosome aberrations either decreased (33----40.5 degrees) or increased (40.5----33 degrees) as early as on day 2, and these patterns were stabilized at the level corresponding to the new conditions. We assayed the induction of DNA breaks in cells, grown at the permissive or nonpermissive temperatures, by using DNA sedimentation in the alkaline sucrose gradient. The DNA sedimentation peaks of cells cultured at 37 and 41 degrees coincided, whereas the DNA of cells cultured at 33 degrees was represented by shorter fragments.

[Induction of the replicative synthesis of DNA by SV40 virus T antigen introduced into cells using liposomes]. / Induktsiia replikativnogo sinteza DNK T-antigenom virusa SV40, vvedennogo v kletki s pomoshch'iu liposom.

The role of virus SV40 T-antigen in the induction of cell DNA synthesis during its incorporation into cell liposomes was studied, using monolamellar liposomes obtained by phase reversal with incorporated highly purified T-antigen. Immunofluorescence studies revealed that T-antigen effectively penetrates inside the cells and after 10 hours is accumulated in the nuclei, where its level remains unchanged for 24 hours. Injections of purified T-antigen into the renal cells of serum-starved CV1 monkeys resulted in an almost 10-fold increase in the number of DNA-synthesizing cells 18 hours after the exposure. The same effect was observed during stimulation of a 10% serum culture. Removal of T-antigen from the preparation by specific immunoadsorption eliminated this effect. Centrifugation of cells grown in the presence of bromodeoxyuridine in a CsCl gradient was used to demonstrate the replicative type of cell DNA synthesis during T-antigen induction.

[Interaction of SV40 T-antigen with tumor cell chromatin]. / Vzaimodeistvie T-antigena virusa SV40 s khromatinom opukholevykh kletok.

The T-antigen of SV40 virus can be found in purified chromatin prepared from virus-induced tumour cells of the Syrian hamster. After treatment of chromatin or isolated nuclei with micrococcal nuclease this protein is detected in the high molecular weight and oligonucleosomal fractions. Data from sedimentation analysis and gel electrophoresis suggest that the T-antigen is predominantly linked with the oligonucleosomal fraction and in a lesser degree with mononucleasomes containing linker DNA and histone H1. A small amount of the T-antigen is found in the mononucleosome complex devoid of histone H1; however, the ratio of the T-antigen to DNA in this case is about 30 times less than that in the oligonucleosomal fraction. In order to investigate the nature of T-antigen binding to nucleosomes, the interaction between the T-antigen and nucleosomes from normal rat liver was studied under restricted binding of the antigen to DNA (pH 8.0). The T-antigen was effectively bound to the nucleosomes and coprecipitated with them in 5 mM MgCl2. It was shown that the T-antigen was adsorbed on columns packed with immobilized histones H1 and nucleosomal histones without H1; the former eluted at 0.15 - 0.25 M NaCl, the latter - at 0.35 - 0.5 M NaCl. The possibility of T-antigen interaction with cellular DNA and protein components of chromatin (primarily to H1) is discussed.

[Interaction of SV40 T-antigen with homologous and heterologous DNA]. / Vzaimodeistvie T-antigena virusa SV-40 s gomologichnoi i geterologichnymi DNK.

Using the DNA filter binding assay, the effects of ionic strength and pH on SV40 T-antigen interaction with viral DNA were studied. The apparent association constants for T-antigen binding to SV40 DNA in Scatchard coordinates in the presence of 40 mM NaCl are equal to 0.67 . 10(6) M-1 (pH 6.0) and 0.86 x 10(7) M-1 (pH 7.4). These data indicate that the interaction between T-antigen and SV40 DNA is more specific at pH 7.4. The coincident values of association constants for T-antigen binding to viral and cellular DNAs (Ka = 0.9 x 10(7) M-1 for cellular DNA) at pH 7.4 and the absence of competition between the two DNA species upon binding with T-antigen suggest that viral and cellular DNAs possess similar sites for T-antigen binding. Denatured DNA competes with viral DNA only at pH 6.0, when the T-antigen--SV40 DNA interaction is less specific.

[Existence of a form of SV40 T antigen incapable of interacting with DNA]. / O sushchestvovanii formy T-antigens virusa SV40, ne sposobnoi Vzaimodeistvovat' s DNK.

The interaction of SV40 T-antigen and viral DNA was studied by using adsorption of DNA-protein complexes on nitrocellulose filters. The T-antigen purification procedure included ion-exchange chromatography on DEAE-cellulose, selective adsorption of cellular proteins on single-stranded DNA-cellulose, chromatography on heparin-Sepharose and removal of cell proteins by an immunosorbent. Only the latter step allowed to remove the contamination of cellular DNA-binding proteins, judging from the reaction of T-antigen neutralization by specific antibodies. It was shown that T-antigen and cellular DNA-binding proteins interact with SV40 DNA at different values of pH, namely ah 6,0-6,4 and 7,9, respectively. The T-antigen obtained was passed through a column with native DNA-cellulose at pH and ionic strength values optimal for interaction with DNA. The bulk of T-antigen (30-40%) did not bind to native thymus DNA and did not interact with SV40 DNA. It is assumed that this fraction is a form of T-antigen, which undergoes structural or functional changes during specific interaction with viral or cellular DNAs.

[Possible role of SV40 T antigen in cell transformation]. / Vozmozhnaia rol' T-antigena Virusa SV40 v transformatsii kletki.

The effect of purified SV40 T antigen on DNA synthesis in isolated nuclei from the confluent culture of CV-1 cells was studied. In the presence of T antigen the incorporation of [3H]TTP into DNA was found to be 2 to 3 times as high as in the control nuclei. The resulting labelled DNA was subjected to alkaline sucrose gradient centrifugation, which revealed the presence of 4S DNA species, corresponding to Okazaki fragments of animal cells. The latter finding suggests a replicative mode of DNA synthesis induced by T antigen. T antigen isolated from the cells infected with SV40 tsA-mutant and kept at a nonpermissive (41 degrees) temperature fails to stimulate DNA synthesis in isolated nuclei from resting cells. On storage at 4 degrees SV40 T antigen gradually loses its ability to stimulate DNA synthesis and by the 8th day even suppresses it when tested on isolated nuclei from a growing cell culture. No effect of T antigen on the endonuclease-induced reparative synthesis of DNA could be observed. The data described suggest that T antigen is directly involved in the control of DNA synthesis in the cells infected or transformed with SV40.

Somatic hybridization and oncogenesis; (Mechanism of formation of malignant tumors and metastases by the action of antilymphocytic serum).

The results of experiments carried out to test some of the consequences of the earlier general theory of oncogenesis, according to which the malignant tumor cell can arise as a result of somatic hybridization of cells of different organ- and tissue-specificity, are described. In the first series a tumor induced by cellophane film, was grafted into syngeneic and allogeneic mice, and antilymphocytic serum (ALS) was then injected. Metastases occurred only in allogeneic recipients receiving ALS. It was thus shown that the ability of cells of this particular tumor to metastasize is not a property inherent in its cells but is acquired by them as a result of interaction with the recipient organism. In the second series it was shown by two immunological methods that the cells of metastases arising under these conditions contain tissue compatibility antigens of donor and recipient origin, i. e., that they are somatic hybridsmin the third series skin from individuals of another strain was grafted on to mice and ALS was injected; hepatomas developed in 74% of these mice. The theory is used to explain several phenomena of carcinogenesis not explicable by other theories: the phenotypic nature of cell transformation, the causes and nature of the duration of the latent period of tumor development, the mechanism responsible for the ability of tumors to overcome the system of immunological defense, the mechanism of activation of endogeneous oncogenic viruses, etc. Finally an answer is given to the question: what is a tumor?

[The potential nuclease activity of the T antigen of SV-40 virus]. / O vozmozhnoi nukleaznoi aktivnosti T-antigena virusa SV-40

SV40 T-antigens were isolated from an extract of golden hamster tumours by precipitation with ammonium sulphate with subsequent fractionation on DEAE cellulose. The degree of purification of the preparation proved to be about 100-fold; it, however, contained an admixture of several cell proteins. Treatment of the DNA of the calf thymus with the T-antigen preparation in the presence of magnesium ions decreased the viscosity of the DNA solution during the first hour of incubation. T-antigen inactivated by heating, and also a fraction of normal hamster tissues analogous to it produced no such effect. In case of centrifugation in the saccharose gradient the constant of DNA sedimentation fell after the treatment with T-antigen from 285 to 165, this corresponding to about4--5-fold reduction of molecular weight of the DNA. The data obtained indicated that the partially purified T-antigen preparation possessed endonuclease activity.