Characterization of an SV40-transformed 3T3 cell line expressing an unusual phenotype

1975 ◽  
Vol 18 (3) ◽  
pp. 427-440
Author(s):  
J.E. Thompson ◽  
J.D. Elligsen ◽  
H.E. Frey

A transformed variant derived as a clone from normal 3T3 cells infected with simian virus 40 (SV40) has been found to possess a phenotype intermediate between that of normal cells and that characteristic of the transformed state, yet cells of the variant still test positively for the SV40-specific nuclear T-antigen. The variant exercises growth control, although not as stringently as do normal cells. Its cell size more closely resembles that of normal cells than of transformed cells. The variant also exhibits levels of spontaneous agglutination that are in line with those characteristic of the normal cells from which it was derived, and far higher than corresponding values for cells exhibiting the fully transformed phenotype. Plasma membranes of variant cells more closely resemble those of transformed cells than of normal cells as estimated by polyacrylamide gel electrophoresis. Perhaps the most distinguishing characteristic of the transformed variant is its complete immunity to agglutination by concanavalin A (Con A), even at concentrations of the lectin as high as 500 mug/ml. Moreover, trypsinization does not render variant cells as agglutinable in the presence of Con A as are untreated fully transformed cells. By contrast the variant displays a low tolerance of Con A toxicity, as monitored by ability to grow after treatment with the lectin, and on this count resembles transformed cells. Moreover a survey of several normal cell lines has revealed that even they do not consistently show resistance to Con A toxicity. These observations indicate that Con A-mediated agglutination and inability to grow after treatment with Con A are quite independent and do not bear a cause and effect relationship.

1985 ◽  
Vol 5 (5) ◽  
pp. 1051-1057
Author(s):  
M Santos ◽  
J S Butel

Cell growth control appears to be drastically altered as a consequence of transformation. Because the cell surface appears to have a role in modulating cell growth and simian virus 40 (SV40)-transformed cells express large T antigen (T-Ag) in the plasma membrane, we investigated whether surface T-Ag expression varies according to cell growth rate. Different growth states were obtained by various combinations of seeding density, serum concentration, and temperature, and cell cycle distributions were determined by flow microcytofluorometry. Actively dividing SV40-transformed mouse cell cultures were consistently found to express higher levels of surface T-Ag and T-Ag/p53 complex than cultures in which cells were mostly resting. In addition, the T-Ag/p53 complex disappeared from the surface of tsA7-transformed cells cultured under restrictive conditions known to induce complete growth arrest (39.5 degrees C), although the surface complex did not disappear from other tsA transformants able to keep cycling at 39.5 degrees C. These results suggest that surface SV40 T-Ag or surface T-Ag/p53 complex, or both, are involved in determining the growth characteristics of SV40-transformed cells.


1985 ◽  
Vol 5 (5) ◽  
pp. 1051-1057 ◽  
Author(s):  
M Santos ◽  
J S Butel

Cell growth control appears to be drastically altered as a consequence of transformation. Because the cell surface appears to have a role in modulating cell growth and simian virus 40 (SV40)-transformed cells express large T antigen (T-Ag) in the plasma membrane, we investigated whether surface T-Ag expression varies according to cell growth rate. Different growth states were obtained by various combinations of seeding density, serum concentration, and temperature, and cell cycle distributions were determined by flow microcytofluorometry. Actively dividing SV40-transformed mouse cell cultures were consistently found to express higher levels of surface T-Ag and T-Ag/p53 complex than cultures in which cells were mostly resting. In addition, the T-Ag/p53 complex disappeared from the surface of tsA7-transformed cells cultured under restrictive conditions known to induce complete growth arrest (39.5 degrees C), although the surface complex did not disappear from other tsA transformants able to keep cycling at 39.5 degrees C. These results suggest that surface SV40 T-Ag or surface T-Ag/p53 complex, or both, are involved in determining the growth characteristics of SV40-transformed cells.


1987 ◽  
Vol 7 (4) ◽  
pp. 1541-1544
Author(s):  
O M Pereira-Smith ◽  
J R Smith

Simian virus 40-transformed human cells fused with other independently derived simian virus 40-transformed cells and tumor-derived cells containing activated H-ras and N-ras oncogenes yielded hybrids capable of indefinite division. Fusions with various other immortal cells yielded hybrids that had limited division potential. T antigen expressed in limited-division hybrids was functional for the induction of cellular DNA synthesis.


1979 ◽  
Vol 178 (3) ◽  
pp. 559-568 ◽  
Author(s):  
E J Niesor ◽  
C B Wollheim ◽  
D H Mintz ◽  
B Blondel ◽  
A E Renold ◽  
...  

The feasibility of infection and transformation by SV40 (simian virus 40) of primary cell cultures derived from newborn-rat pancreas was investigated. As judged by the presence of intranuclear SV40 T-antigen, exposure to the virus resulted specifically in infection and transformation of epithelioid (predominantly endocrine) cells. The transformed cells were subcultured (more than 64 passages) and cloned. Culture medium and acid/ethanol extracts of the cells did not contain detectable amounts of immunoreactive insulin after the third subculture. However, inoculation of such SV40-transformed pancreatic cells into immunodeficient rats results in tumours in which insulin production was partially restored through the passage in vivo, since the tumour cells contained and synthesized small amounts of immunoreactive insulin which co-migrated with an insulin marker on gel chromatography. Interestingly, the transformed cells maintained under tissue-culture conditions produced a protein immunologically related to insulin, soluble in aqueous buffer but insoluble in acid/ethanol. This 3000-dalton protein is too large to be a translation product of the rat preproinsulin 9S mRNA. SV40-transformed pancreatic cells might prove useful in the investigation of the factors controlling and maintaining insulin biosynthesis.


1979 ◽  
Vol 29 (1) ◽  
pp. 69-75 ◽  
Author(s):  
C Chang ◽  
R G Martin ◽  
D M Livingston ◽  
S W Luborsky ◽  
C P Hu ◽  
...  

The transforming protein of Simian virus 40 (SV40), large T-antigen, regulates transcription both positively and negatively during the productive infection cycle. We have isolated a number of cellular genes which are expressed at elevated levels in SV40-transformed cells and have used these to study the mechanism or mechanisms by which the viral transforming protein regulates cellular gene expression. Small RNAs homologous to the mouse B2 repetitive sequence family are found at higher levels in transformed cells than in normal cells and we have shown that pure large T-antigen stimulates transcription of such repeats by RNA polymerase III. A class I major histocompatibility complex (MHC) gene is also activated as a result of SV40 transformation and we have used DNA-mediated gene transfer to study how this gene is regulated by large T-antigen.


1984 ◽  
Vol 4 (10) ◽  
pp. 2180-2186
Author(s):  
O Pinhasi ◽  
M Oren

DNA specific for the murine p53 cellular tumor antigen was linked to the early simian virus 40 promoter and introduced into monkey COS cells either by transfection with recombinant plasmids or by infection with virus. Recipient cells made substantial amounts of a protein apparently identical to mouse p53. Severalfold-larger quantities were detected when cells were transfected with an intron-containing p53-specific segment, as compared with transfection with intronless cDNA. The p53 encoded by the recombinant DNA was capable of complexing with the simian virus 40 T antigen. Transfected p53 was also probably associated with a cellular 68-kilodalton protein, which may be related to a protein coprecipitating with p53 in some transformed cells. These findings confirm the predicted reading frame and protein boundaries and demonstrate that apparently functional p53 can be produced in cells via experimentally introduced recombinant DNA.


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