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Biomedical subjects

J T August

Publications and source records attributed to J T August.

At least 19 recordsLinked to original sources
PubMed1978

Transport and phosphorylation of hexoses in normal and Rous sarcoma virus-transformed chick embryo fibroblasts.

Effects of transformation by Rous sarcoma virus on sugar uptake and activity and the subcellular distribution of hexokinase isozymes in chick embryo fibroblasts were examined. Transformation caused a several-fold increase in the maximum velocity for uptake of 2-deoxyglucose without a significant change in Km. Cytochalasin B (CB), was used to differentiate between the effects of transformation on facilitated diffusion and the nonsaturable (CB-insensitive) mode. Transformation was found to stimulate 2-deoxyglucose transport by both mechanisms, but the increase in transport by the CB-insensitive mode was greater. Transformation enhances the activity of hexokinase, the enhancement being confined to the particulate fraction of the enzyme. Heat-inactivation and electrophoretic mobility studies showed that although hexokinase Type I is the major form in both normal and transformed fibroblasts, there is a significant increase in the proportion of the Type II isozyme in the transformed cells.

Animals↗
PubMed1978

Polypeptide maps of cells infected with murine type C leukemia or sarcoma oncovirus.

The polypeptide composition of murine fibroblast cells and the effect of infection by RNA sarcoma and leukemia viruses were analyzed by two-dimensional gel electrophoresis and tryptic peptide mapping. The polypeptide maps of NIH Swiss mouse embryo fibroblasts (NIH/3T3) and BALB/c mouse embryo fibroblasts (BALB/3T3) were very similar except for two major polypeptides of about 65,000 and 75,000 daltons which were not detected in BALB/3T3 cells. NIH/3T3 cells infected with either Rauscher or Gross oncoviruses and outbred Swiss mouse embryo fibroblasts (3T3 FL) showed two major polypeptrides of about 73,000 and 80,000 daltons not found in uninfected NIH/3T3 cells. The 3T3 FL cells, although uninfected, were also found to contain a high concentration of envelope glycoprotein of an endogenous oncovirus. 3T3 FL cells transformed by Moloney sarcoma virus showed changes in many polypeptides, including several major components: the disappearance or modification of a component of 60,000 daltons, an increased concentration and shift in pl of a glycoprotein of 48,000 daltons, and the apparent loss of several smaller polypeptides. None of the major changes of the transformed cells were associated with cell surface proteins labeled by lactoperoxidase-catalyzed iodination.

AKR murine leukemia virus↗
PubMed1978

Further evidence for deletion of envelope glycoprotein (gp69/71) sequences in formation of Moloney-murine sarcoma virus.

Moloney-murine sarcoma virus (S+L- strain of M-MSV) has been nonproductively cloned in murine and non-murine host cells (S+L- cells) and the expression of Moloney leukaemia virus (M-MuLV) 30000 mol. wt. core protein (p30) and envelope glycoprotein (gp69/71) were studied by radioimmunoassay. Antigenic determinants of the M-MuLV p30 were associated with the sarcoma virus genome in these non-productively transformed cell clones studied, while the determinants of M-MuLV gp69/71 were not. The absence of envelope-associated glycoprotein expression in sarcoma virus transformed cells was confirmation of biological studies demonstrating that rescued sarcoma virions acquire envelope-associated properties of host range, neutralization and interference from rescuing helper virus, and further evidence that the M-MuLV gp69/71 sequences have been deleted during the formation of the M-MSV. During the course of these studies, it was also found that S+L- dog cells were releasing into culture supernatant large amounts of the p30 antigenic determinant, apparently as a soluble antigen.

Cell Line↗
PubMed1978

Stoichiometry and specificity of binding of Rauscher oncovirus 10,000-dalton (p10) structural protein to nucleic acids.

A structural protein of Rauscher oncovirus of about 8,000 to 10,000 daltons (p10), encoded by the gag gene, has been purified in high yield to apparent homogeneity by a simple three-step procedure. The purified protein was highly basic, with an isoelectric point of more than 9.0, and its immunological antigenicity was chiefly group specific. A distinctive property of the protein was the binding to nucleic acids. The stoichiometry of p10 binding to Rauscher virus RNA was analyzed using both 125I-labeled p10 and 3H-labeled RNA. The protein-RNA complex, cross-linked by formaldehyde, was separated from free RNA and free protein by velocity sedimentation and density gradient centrifugation. A maximum of about 140 mol of p10 was bound per mol of 35S RNA, or about one molecule of p10 per 70 nucleotides. This protein-RNA complex banded at a density of about 1.55 g/ml. The number of nucleic acid sites bound and the affinity of p10 binding differed significantly among the other polynucleotides tested. The protein bound to both RNA and DNA with a preference for single-stranded molecules. Rauscher virus RNA and single-stranded phage fd DNA contained the highest number of binding sites. Binding to fd DNA was saturated with about 30 mol of p10 per mol of fd DNA, an average of about one p10 molecule per 180 nucleotides. The apparent binding constant was 7.3 X 10(7) M(-1). The properties of the p10 place it in a category with other nucleic acid binding proteins that achieve a greater binding density on single-stranded than on double-stranded molecules and appear to act by facilitating changes in polynucleotide conformation.

Binding Sites↗
PubMed1977

Polypeptides of cells transformed by RNA or DNA tumor viruses.

Proteins solubilized from normal BALB/3T3 cells and BALB/3T3 transformed by simian virus 40 or Kirsten sarcoma virus have been analyzed by two-dimensional gel electrophoresis and tryptic peptide mapping. A large fraction of the polypeptides of the virus-transformed cells, about 30%, were different from normal cells. In contrast to the marked differences between normal and transformed cells, the polypeptides of the DNA and RNA virus-transformed cells were almost identical. These findings were observed with polypeptides stained by Coomassie Blue, or labeled with [14C]glucosamine or [35S]methionine. Pulse-chase analysis showed that most of the polypeptides were stable during 20 hr of incubation. The identity of several polypeptides was confirmed by tryptic peptide mapping.

Cell Line↗
PubMed1977

Biochemical and immunological characterization of the major envelope glycoprotein gp69/71 and degradation fragments from Rauscher leukemia virus.

Analysis of the proteins of Rauscher murine oncornavirus by immunoprecipitation showed that antiserum to the purified envelope glycoprotein of approximately 69,000 and 71,000 daltons (gp69/71) reacted as well with a number of other components of several murine oncornaviruses of approximately 45,000, 32,000, and 15,000 daltons. Polypeptides of similar size were also produced by limited proteolysis of purified gp69/71; these degradation fragments were shown to contain carbohydrate by the incorporation of (3)H from sodium boro[(3)H]hydride after neuraminidase and galactose oxidase treatment. Each of these glycoproteins was isolated by preparative polyacrylamide gel electrophoresis and was analyzed by tryptic peptide mapping. The major virion components of 69,000 and 71,000 daltons were nearly identical, as were the primary degradation fragments. Analysis of the immunological properties of the glycoproteins showed that the 71,000-, 69,000-, and 32,000-dalton glycoproteins behaved similarly with respect to type and group-specific antigenic determinants. In contrast, the 45,000-dalton glycoprotein lacked detectable interspecies and some of the group-specific reactivity. Components of about 45,000 and 32,000 daltons isolated directly from virions were also identified as constituents of the major envelope glycoprotein by immune precipitation and tryptic peptide mapping. These results indicate that all of the examined virion glycoproteins of approximately 71,000, 69,000, 45,000, and 32,000 daltons are derived from the same viral gene and that these lower-molecular-weight glycoproteins can readily be produced from the major envelope glycoprotein.

Carbohydrates↗
PubMed1977

Murine Type C viral envelope glycoprotein gp69/71 and lupus-like glomerulonephritis of New Zealand mice. An immunoperoxidase study.

The location of murine Type C viral envelope glycoprotein antigen in the glomerulonephritic kidneys of NZB and NZB/NZW F1 hybrid mice was analyzed by the indirect immunoperoxidase technique at light and electron microscopic levels. Immune complex deposits of the glycoprotein antigen were present in the glomeruli in mesangial and subepithelial sites. In addition to the glomerular depositions, viral envelope antigen was also present at the brush border of proximal tubular epithelial cells and in lymphoid cells infiltrating the kidneys of these mice.

Animals↗
PubMed1977

Characterization of tumour virus proteins. I. Radioimmunoassay of the P27 protein of avian viruses.

The major structural protein of avian oncornaviruses, a core component of about 27000 daltons, has been measured by radioimmunoassay. The purified protein was labelled with 125Iodine by chloramine-T method. The immune serum titer was defined as the highest serum dilution able to precipitate 50% of the labelled antigen present in the system. Standard competition curve was constructed in order to determine the equivalents of protein, in a system with limiting antibody concentration. In the experimental conditions used, 0.14 ng of AMV-P27 inhibited 50% of 125I-AMV-P27 (1.0 ng) precipitation. The 125I-AMV-P27 vs anti-AMV-P27 system was used to study the competition of normal cells, purified virus suspension, productive cells and supernatant fluids. Most of the chicken embryo fibroblasts showed expression of this viral component. The phenomena of cell transformation, the increase in total protein, and the expression of P27 were studied in rapid transformation of CEF by RSV-SRA.

Alpharetrovirus↗
PubMed1977

Characterization of tumour virus proteins. II. Expression of the protein P30 in transformed productive and non-productive Ki/NRK cells.

The structural protein of murine tumour virus P30 has been measured by radioimmunoassay. The titer of each serum was determined by using as antigen the purified Rauscher viral protein labeled with 125iodine. Standard competition curve was constructed in order to determine the equivalent of protein to inhibit the precipitation reaction under limited antibody concentration. Competition by purifed Kirsten virus suspension, normal rat kidney cells, transformed-productive and transformed non-productive cells were measured in homologous and heterologous system. Type, group and interspecies determinants were characterized using the proper antigen-antibody system. Once the proteins have interspecie determinants, it is possible that we might be able to use some mammalian virus protein as tool to determine the presence of viral protein in human processes.

Animals↗
PubMed1976

Characterization of a virion protein kinase as a virus-specified enzyme.

Antibodies which completely inhibited the enzymatic activity of the protein kinase associated with virions of frog virus were obtained by immunization of rabbits with the purified enzyme. This inhibition provided a specific probe for the frog virus protein kinase, since this antiserum had no inhibitory effect on a variety of other protein kinases, including the activity of uninfected cells, or the protein kinase associated with vesicular stomatitis virus or vaccinia virus cultivated in the same cell line as frog virus. The frog virus protein kinase was characterized as a virus-specified protein on the basis of the following observations: (a) the virion protein kinase was antigenically distinct from essentially all of the protein kinase expressed in uninfected cells; (b) following infection by frog virus more than a 15-fold increase was detected in the specific activity of intracellular protein kinase and most of this activity was antigenically related to the virion enzyme; (c) when frog virus was grown in cells derived from widely different species, the antigenic and biochemical specificities of the virion protein kinase remained identical; and (d) screening of cells infected with different temperature-sensitive mutants of frog virus indicated that certain viral mutants failed to synthesize this protein kinase when cultivated at the nonpermissive temperature.

Adenosine Triphosphate↗