The surprising finding of Huebner et al.1 that tumors developing in hamsters after inoculation with adenovirus 7 contain the complement-fixing cellular antigen2 of SV40 transformed cells suggested that preparations of adenovirus 7 should -be studied for ability to induce similar antigens in cells growing in culture. Recent observations using the immunofluorescence technique had confirmed that cells transfotmed by SV40 contain a new cellular antigen; this antigen appears to be synthesized in the nucleus and is present in all cells of cultures derived from SV40-transformed systems.3' 4 These sensitive techniques have made it possible to detect the tumor antigens even when they are synthesized in relatively few cells in the culture. The same techniques have been used to demonstrate that the SV40 tumor antigens occur not only in virus-free transformed cells but are also synthesized in cells infected with SV40 prior to the appearance of the virus antigens.5 6 This report will document the finding that preparations of adenovirus 7 grown in monkey cells and free of infectious SV40 can induce the synthesis of a new intranuclear antigen unrelated to adenovirus in a wide variety of cells. This antigen is similar to or identical with the tumor antigens induced by SV40 virus. These results suggest the incorporation and transmission of a portion of the SV40 genome by the unrelated adenovirus.Materials and Methods.-Viruses: The L. L. strain of adenovirus 7 used in these studies, kindly made available by Wyeth Laboratories, was originally isolated and passed serially in primary rhesus kidney cells. Stocks of the virus were subsequently found to be contaminated with SV40 and were therefore treated with SV40 antiserum and passed twice in primary green monkey kidney (GMK) cells in the presence of the antiserum. The virus was then passed 4 times in the absence of antiserum in GMK cells; large volumes of each virus yield were tested and found to be free of infectious SV40. This virus stock was designated E46. A stock of virus consisting of several liters was prepared by passing E46 in GMK, and the harvest was designated SP2. Samples of lots E46 and SP2 were kindly supplied by Dr. B. A. Rubin of Wyeth Laboratories. Repeated attempts to isolate SV40 from E46 and SP2 have been unsuccessful. Complement fixation tests with SV40 antiserum have also failed to detect the presence of SV40 antigen. A fresh isolate of adenovirus 7 from a fatal human case7 passed only in human embryonic kidney (HEK) cells was also employed.Cells: The various cells utilized were grown in Eagle's basal medium fortified by varying concentrations of calf serum ranging from 2 to 10%; GMK cells were grown in Melnick's medium.Immunofluorescence tests: The cells were trypsinized and grown on 15-mm cover glasses in 60-mm plastic Petri dishes in 5% CO2 at 37°C. When monolayers had formed, 0.1 ml of virus inoculum was added per cover glass, adsorption allowed to occur for 1 hr at 37°C, and 5 ml of maintenance fluid added per Petri dish. The cultures were reincubated at 37...

No abstract

The demonstration that hamster (1-3) and human (2) cells transformed by SV40 virus synthesize a new antigen under control of the virus genome led to the discovery that the same antigen is also formed during the normal replication cycle of SV40 in green monkey cells (3-5). As the sequence of SV40 replication had been previously studied (6, 7), it was quickly ascertained that this new tumor or T antigen is formed during the latent period and that synthesis of T antigen precedes that of the virus or V antigen found in the SV40 virion (5).From our earlier investigations of the inhibitory effect of 5-fluorouracil and 5-fluorodeoxyuridine on the synthesis of V antigen and infectious virus (7), we were led into a study of the effects of these and other DNA antagonists and inhibitors on the synthesis of both T and V antigens in an effort to obtain information concerning the nature and role of the T antigen in the synthesis of SV40 virus as well as its formation in cells transformed by the virus but free of infectious virions. Materials and MethodsViru*.--Stocks of SV40 were prepared in primary African green monkey kidney cells (GMK) growing in 16-ounce bottles. When cytopathic effects (CPE) involved 75 to 100 per cent of the cells, the cultures were disrupted by quick-freezing and thawing. After clarification by centrifugation at 3000 g for 5 minutes at 4°C, the virus was dispensed in ampoules which were sealed; the virus was quickly frozen, and stored at --90°C. "liters of the virus stocks were obtained using the plaque assay previously described (8); all stocks employed in this study contained 108 to 10 T plaque-forming units (PFU) per ml. The tumorigenic potential of this virus strain has been reported from this laboratory (9), as has the growth cycle of the virus in GMK cells (7).

No abstract

Cells infected with the papovavirus SV40 not only synthesize viral antigen but also synthesize the specific nonviral antigen found in SV40-induced tumors. In the presence of the DNA antagonist cytosine arabinoside, infected cells fail to make viral antigen but still synthesize the tumor antigen. Iododeoxyuridine does not inhibit the synthesis either of tumor or of virus antigen but does prevent the development of infectious virus.