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S W Luborsky

Publications and source records attributed to S W Luborsky.

At least 19 recordsLinked to original sources
PubMed1981

Surface proteins of simian-virus-40-transformed cells.

Mammalian cells transformed in tissue culture by SV40 were shown to contain, in addition to the SV40-coded 94,000 d large T antigen and the 20,000 d small t antigen, a approximately 56,000 d cellular protein, which specifically precipitates with sera of animals bearing SV40-induced tumor(s) (tumor or T serum). We investigated the presence of these three proteins at the surface of logarithmically growing SV40-transformed cloned mouse cells, after metabolic labelling with [35S]-methionine for 3 h. The 56,000 d protein was found to be susceptible to digestion by trypsin under conditions which did not disrupt the cells, while no small t antigen was found to be digested. Both the 56,000 d cellular protein and the SV40 large T antigen were susceptible to lactoperoxidase-catalyzed iodination from the outside of intact cells. Trypsin treatment removed both the iodinated 56,000 d protein and the iodinated SV40 large T antigen. These experiments indicated that (a certain amount of) the 56,000 d protein and a relatively small amount of the large T antigen (which is present mainly in the nucleus) are present on the cell surface. The results confirm and extend independent experiments using subcellular fractionation techniques (Luborsky and Chandrasekaran, 1980; Soule and Butel, 1979). After heat treatment (at 50 degrees C for 30 min) of the whole-cell extract the 56,000 d cellular protein was precipitated by the tumor serum in the absence of precipitation of SV40 large T antigen. This result showed that the 56,000 d protein is more (thermo)stable (in the whole-cell extract) than the SV40 large T antigen, and also indicated that the tumor serum employed had antibodies against the 56,000 d cellular antigen. The heat-treated whole-cell extract of Sv40-transformed mouse cells was able to immunize and fully protect mice against a lethal tumorigenic dose of SV40-transformed cells. These results suggest the need for further experiments to characterize the chemical and immunologic properties of the 56,000 d protein.

Animals↗
PubMed1980

Subcellular distribution of simian virus 40 T antigen species in various cell lines: the 56K protein.

The subcellular distribution of SV40 anti-T serum-specific species was examined in SV40-transformed, T-antigen-positive tissue culture cell lines of rat and of AL/N and BALB/c mouse origin. Cells were labelled with [35S]methionine. The cytoplasm, nuclear and membrane fractions were obtained, and their radioimmunoprecipitates analyzed by gel electrophoresis. Tests were performed to determine the purity of these subcellular fractions, and negligible cross-contamination was found. The cytoplasm fractions lacked detectable anti-T serum reactivity. Large amounts of both large T antigen and a 56K protein were always present together both in the nuclear fractions and, in a somewhat lesser amount, in the plasma membrane fractions of all cell lines examined. Analysis of density gradient sedimentation profiles of the immunoprecipitates of whole-cell extracts indicated these species were associated in some fashion, probably with each other. The activity of the 56K protein may be associated with its presence on the cell surface where, either alone or acting together with the large T antigen, it might provide the surface activity responsible for tumor-specific surface and/or transplantation antigen activities.

Animals↗
PubMed1979

Relationship between T-antigen and tumor-specific transplantation antigen in simian virus 40-transformed cells.

The simian virus 40 (sv40) tumor antigen (T-antigen) and tumor-specific transplantation antigen (TSTA) have been partially purified and studied to clarify their relationship. The T-antigen and the TSTA were partially purified from nuclei of SV AL/N cells, and SV40-transformed mouse embryo fibroblast line, by precipitation with ammonium sulfate and chromatography on DEAE- and DNA-cellulose. The T-antigen was assayed by complement fixation, and the TSTA was assayed by its ability to immunize mice against SV40-containing ascites tumor cells. When T-antigen- and TSTA-containing preparations were sedimented through sucrose gradients, each antigen had a major peak of activity at a sedimentation coefficient of 6.7 and minor peaks in other regions. Antiserum against T-antigen (from tumor-bearing hamsters) immunoprecipitated the TSTA activity. A preparation of T-antigen from human SV80 cells, which exhibited only one protein band after sodium dodecylsulfate-polyacrylamide gel electrophoresis, had TSTA activity when as little as 0.6 microgram of protein per mouse was used for immunization. These experiments demonstrate that the T-antigen, the product of the SV40 early A gene is capable of inducing specific immunity against transplantation of SV40-transformed tumor cells in mice.

Antigens, Neoplasm↗
PubMed1978

Comparative behavior of simian virus 40 T-antigen and of tumor-specific surface and transplantation antigens during partial purification.

The simian virus 40-specific T-antigen has been extracted from SV AL/N mouse embryo tissue culture cells by treatment with Triton X-100 detergent. The extracts contained tumor-specific transplantation antigen (TSTA) and tumor-specific surface antigen. These extracts were purified by ammonium sulfate precipitation and diethyl-aminoethyl cellulose and phosphocellulose column chromatography and were assayed for the three antigens. We found that T-antigen, TSTA, and much of the tumor-specific surface antigen copurified through all purification steps. This finding is consistent with previous suggestions of the close degree of homology that must exist between the protein species carrying these three antigenic determinants. The antibody-mediated cytolytic assay appears to detect a new type of antigen on the cell surface, different from T-antigen and TSTA; two antigenic fractions were obtained from the phosphocellulose column that had tumor-specific surface antigen activity, but one of these did not have T-antigen or TSTA activities.

Animals↗
PubMed1977

Metabolically labeled cell membrane proteins in spontaneously and in SV40 virus transformed mouse fibroblasts.

A family of mouse fibroblast cell lines in exponential phase of growth were compared in protein constitution of their cell membranes. In preparations from these cells enriched in cell-surface membrane we observed one protein component (apparent molecular weight about 250 000) consistently to be reduced or absent in an SV40 virus transformed cell line, when compared with the normal cell line. No such compositional difference was observed in a spontaneously transformed tumorigenic clonal derivative cell line, or in subclones of such a derivative cell line, with or without SV40 virus infection. However, in metabolic labeling experiments with 14C-labeled mixed amino acids, a consistent decrease also was demonstrated in the biosynthesis of the same protein in the SV40 virus infected subclone, as compared to an uninfected sister subclone, during exponential growth. This specific difference in biosynthesis is apparently related to the presence and functioning of the SV40 gene, and correlates with the ability of these cells to grow in viscous medium, but not with cellular tumorigenicity.

Animals↗
PubMed1977

Detergent solubilization and partial purification of tumor specific surface and transplantation antigens from SV40-virus-transformed mouse cells.

A solubilization technique employing 0.5% Triton X-100 was developed to obtain both SV40 virus (SV40)-induced tumor-specific surface antigen(s) (TSSA) from SV40-transformed mouse cells, as determined by a serum-mediated microcytolytic assay, and tumor-specific transplantation antigen(s) (TSTA), as determined by in invivo experiments. High yields (approximately 50%) of TSSA were obtained in whole-cell extracts and also after ammonium sulfate fractionation. Additional fractionation of a 30-50% ammonium sulfate fraction by gel exclusion chromatography on Sephadex G-150 resulted in two pooled fractions which contained TSSA activity. The first eluted close to the void volume, and the second in the 45,000 molecular weight region. The various TSSA active fractions were also active in vivo TSTA tests. Detergent solubilization provides a suitable technique to recover the SV40-induced antigens in good yield, and apparently in intact form.

Antigens, Neoplasm↗