Howard M. Temin

Characteristics of an assay for Rous sarcoma virus and Rous sarcoma cells in tissue culture

Abstract An accurate tissue culture assay for Rous sarcoma virus (RSV) and Rous sarcoma cells is described. The Rous sarcoma virus changes a chick fibroblast into a morphologically new and stable cell type with the same chromosomal complement as ordinary chick embryo cells. One virus particle is enough to change one cell, but at any one time 90% of the cells in a culture are not affected by RSV. The cellular resistance is the same in a clonal population. The physiological state of the cell is of some importance in deciding whether or not it is competent to be infected by RSV but so far attempts to infect all the cells in a chick embryo culture by altering the physiological condition have failed.

Formation of infectious progeny virus after insertion of herpes simplex thymidine kinase gene into DNA of an avian retrovirus

We have prepared several infectious stocks of an avian retrovirus, spleen necrosis virus, containing the herpes simplex virus type 1 thymidine kinase (tk) gene. The viruses were produced after cotransfection of chicken cells with DNA from recombinants between cloned spleen necrosis virus and tk DNAs and DNA of cloned reticuloendotheliosis virus strain A. removal of sequences in the tk gene for the end of tk mRNA increased a thousand fold the yield of infectious recombinant virus. Infection of chicken or rat tk- cells with the recombinant virus transformed them to a tk+ phenotype.

The participation of DNA in Rofs sarcoma virus production

Abstract Experiments are presented showing that the inhibition by actinomycin D of Rofs sarcoma virus production, which was reported previofsly ( Temin, 1963b) , is due to the inhibition of synthesis of viral nucleic acid. The results of experiments on the effects of amethopterin on Rofs sarcoma virus (RSV) production, on the induction of converted nonvirus-producing Rofs cells, and on the infection of normal cells with RSV are presented. There appear to be some steps in infection which are sensitive to inhibition by amethopterin which are not present in virus production or induction.

The control of cellular morphology in embryonic cells infected with rous sarcoma virus in vitro.

Abstract Different morphological changes are induced in chick fibroblasts in vitro by infection with various lines of Rous sarcoma virus. The type of change induced is a genetic character of the virus line. The host cell also plays a role in determining the cellular change, the same virus line producing different effects on different kinds of fibroblasts. Viral mutations occur during the multiplication of the virus in Rous sarcoma cells and not at the time of infection.

The effects of actinomycin D on growth of Rous sarcoma virus in vitro

Abstract Low concentrations of actinomycin D (0.1 μg/ml) reversibly inhibit production of Rous sarcoma virus (RSV) by Rous sarcoma cells. Higher concentrations inhibit irreversibly. Genetically resistant variants of RSV are not found. Infection of cells and initiation of virus production are also inhibited.

A radiological study of cell-virus interaction in the rous sarcoma☆

Abstract Rous sarcoma virus (RSV) and Newcastle disease virus (NDV) are in activated exponentially by X-rays and ultraviolet (UV) light. While both viruses are similar in their sensitivity to X-rays, RSV is about ten times more resistant to UV light than is NDV. The capacity of chick fibroblasts to initiate the growth of RSV is as sensitive to inactivation by both X-rays and UV light as is their ability to divide and form colonies, while their capacity to initiate the growth of NDV is many times more radioresistant. A common radiosensitive target controls the ability of cells to divide and to initiate the production of RSV. Once the cells have started to produce virus, they may upon irradiation lose their ability to divide and yet continue to produce virus. The results suggest that the genome of the Rous sarcoma virus must be integrated with that of the cell before virus production can begin.

Different localization of the product of the v-rel oncogene in chicken fibroblasts and spleen cells correlates with transformation by REV-T

Reticuloendotheliosis virus strain T (REV-T) is a highly oncogenic avian retrovirus that transforms early lymphoid cells in vivo and in vitro, but REV-T does not transform chicken embryo fibroblasts (CEF). Using antisera to p59v-rel, the v-rel oncogene product of REV-T, we show that p59v-rel is expressed at equal levels and is a phosphoprotein in REV-T infected spleen cells and CEF. Biochemical fractionation and immunofluorescence of REV-T infected nontransformed CEF show that p59v-rel is loosely associated with the nucleus. However, in REV-T transformed spleen cells p59v-rel is primarily a cytoplasmic protein. MSB-1 cells, a Mareks disease virus transformed T cell leukemic line, and E26 virus transformed myeloid cells show nuclear staining of p59v-rel when they are infected by REV-T. Our results indicate that there is a correlation between a cytoplasmic localization of p59v-rel and transformation by REV-T, and they suggest that p59v-rel cannot transform cells in which it assumes solely a nuclear location.

Recent advances in retrovirus vector technology

Retroviral vectors are widely used for the study of retroviral replication and to introduce DNA into somatic cells. An exciting new approach in retroviral vector technology is the use of internal ribosome entry sites from picornaviruses to provide stable expression of multiple genes. In addition, strategies are being developed that target the expression of retroviral vectors to specific cell populations.

Retrovirus Vectors for Gene Transfer: Efficient Integration into and Expression of Exogenous DNA in Vertebrate Cell Genomes

Retroviruses are animal viruses with an RNA genome that replicate through a DNA intermediate. The DNA intermediate, the provirus, is stably integrated into cellular DNA. (Alternatively, retroviruses can be considered the RNA form of cellular movable genetic elements that transpose through an extracellular RNA intermediate.) Because of this efficient integration and other properties to be discussed, retroviruses are considered the most likely vectors for use in human gene therapy (Wyngaarden, 1985).

Malignant Transformation of Cells by Viruses

Most of the present approaches to an answer to the question How do viruses cause cancer? (as opposed to the question Do viruses cause human cancer?) come from studies involving cells in culture. In this article, I shall discuss from this perspective—and in a somewhat personal and speculative fashion—the general nature of cancer, some properties of cancer cells in cell culture, and some results from the study oftumor viruses in cell culture.